In this work, we demonstrate sub-10 nm uniform deposition of Al2O3 on MoS2 basal plane by applying ultra-low energy remote oxygen plasma pretreatment prior to atomic layer deposition. It is demonstrated that oxygen species in ultra-low energy plasma are physically adsorbed on MoS2 surfaces without making the flakes oxidized, and is capable of benefiting the mobility of MoS2 flake. Based on this method, top-gated MoS2 transistor with ultrathin Al2O3 dielectric is fabricated. With 6.6 nm Al2O3 as gate dielectric, the device shows gate leakage about 0.1 pA/μm2 at 4.5 MV/cm which is much lower than previous reports. Besides, the top-gated device shows great on/off ratio of over 108, subthreshold swing (SS) of 101 mV/dec and a mobility of 28 cm2/Vs. With further investigations and careful optimizations, this method can play an important role in future nanoelectronics.The integration of ultra-thin gate oxide, especially at sub-10 nm region, is one of the principle problems in MoS Since it enables the individual control of each device, the realization of high-performance dual gate or top-gated MoS2 transistors is a necessary step towards the practical application181213141516172 surfaces. Even though uniform atomic layer deposition (ALD) process of Al2O3 on MoS2 at 200 °C has been reported by Liu et al.2 may be feasible, other works have shown that with pristine MoS2 flakes the direct deposition of high-κ dielectrics form island-like clusters, owing to the absence of dangling bonds on MoS2 basal plane202O3 at 200 °C in Liu’s work2O3 layer grown on MoS2 flakes. The island-like growth of high-κ dielectrics on MoS2 would induce a large leakage current, therefore top-gated high-κ dielectrics in nearly all the previous reports are very thick and MoS2 transistors with sub-10 nm top gate dielectrics are seldom reported181213141516172 transistors are 50 nm Al2O32et al. and Krasnozhon D. et al., respectively. Only by realizing high-quality pinhole-free and thin dielectrics over large area on MoS2 can the continual scaling down of MoS2 FETs be possible. With the shrink of dielectric thickness, especially at sub-10 nm region, the gate capacitance would be greatly improved, leading to better control of the channel and larger drive current. Some methods have been proposed to achieve uniform growth of high-κ materials on MoS2, such as an ultrathin metal oxide buffer layer, organic functionalization of MoS2 and ultraviolet-ozone exposure2021Excellent performance of MoS2O3 growth on MoS2 basal plane by applying a remote O2 plasma treatment prior to Al2O3 growth is proposed, and top-gated MoS2 MOSFET with ultrathin Al2O3 dielectric deposited using this method is also studied. Notably, the Al2O3 dielectric layer is about 6.6 nm, which is the thinnest top gate dielectric ever reported, but exhibits the impressive leakage current about 0.1 pA/μm2 at 4.5 MV/cm. This leakage is even much smaller than that of MoS2 transistors capped with much thicker top gate dielectrics1813168, subthreshold swing (SS) of 101 mV/dec and a mobility of 28 cm2/Vs. In addition, mechanism investigations show that after the pretreatment, oxygen atoms are physically adsorbed on the MoS2 surface without oxidizing it. This non-destructive physical adsorption mechanism is revealed by the advanced ultra-high-vacuum (UHV) in-situ analysis system. We believe it will benefit the two-dimensional electronic devices research a lot.In this work, a CMOS process compatible method to achieve uniform Al2O3 growth on pristine MoS2, functionalization of the MoS2 surface is required to introduce uniform surface groups that serve as active nucleation sites for the ALD process212 flakes was carried out with a plasma enhanced ALD system. For the sample in 2O3 was directly deposited on MoS2 surface at 200 °C using TMA (Trimethyl Aluminum) and H2O as precursors, which were kept at 18 °C in stainless bottles. By comparison, the sample in 2 plasma treatment before ALD. The pretreatment contained two steps. Each step consisted of 30 s remote O2 plasma exposure followed by purging with Ar for 5 s. Afterwards, 120 cycles Al2O3 was deposited in the same chamber. The different growth topography of Al2O3 on MoS2 basal planes are shown in 2 basal plane, direct deposition of Al2O3 films are in forms of island-like clusters with large pinholes, and the lateral size of most pinholes are over 100 nm. In this situation, it is easy to imagine that the gate dielectric of top gate MoS2 transistors have to be thick to form uniform film which is necessary to keep the leakage current at a sufficient lower level. By contrast, with remote oxygen plasma treatment prior to ALD, the grown Al2O3 film is completely uniform on MoS2 surface as shown in 2 plasma pretreatment served as an effective method to supply sufficient nucleation sites to achieve a uniform ALD process. More details about the direct deposition of Al2O3 on pristine MoS2 basal planes are available in the Supplementary Information. According to previous reports with graphene2252 flakes. Therefore, further analysis is needed to investigate the impact of the low energy remote oxygen plasma.Leakage current of gate oxide results in high power consumption and performance degradation of the two dimensional layered transistorsIn-situ investigations were performed to gain an insight into the mechanisms of uniform growth after a remote O2 plasma pretreatment, looking at whether the MoS2 flakes were oxidized during the treatment or oxygen atoms were adsorbed on the MoS2 surface and acted as nucleation sites in the following ALD process. In the in-situ cluster system, the ALD system was connected to an X-ray photoelectron spectroscopy (XPS) system through a high-vacuum transfer line (the pressure was about 10−10 mbar). The sample was first transferred from the load-lock chamber to the XPS chamber for characterization. It was then transferred to the ALD chamber for 5 s remote O2 plasma treatment. Afterwards, the sample was transferred back into the XPS chamber for further measurements. This procedure was repeated twice with treatment time of 15 and 30 s. The Mo 3d, S 2s and S 2p regions of the XPS spectra are shown in 4+ 3d5/2 and Mo4+ 3d3/2 components, respectively. Similar peaks appear around 161.8 and 163 eV, referring to the S 2p3/2 and S 2p1/2 components of the S 2p region, respectively. In 2 plasma treatment, implying that the chemical bonds were not damaged during the process. Also, no peaks appear around 236 eV, demonstrating that the molybdenum atoms were not oxidized after the plasma treatment2725262 surfaces and acted as nucleation sites for the ALD process. The remote plasma was gentle enough such that the flakes were not oxidized during the treatments. When the flakes were transferred into the XPS chamber through the transfer line, the adsorbed oxygen atoms desorbed, caused by the high vacuum in the transfer line and the reductive environment due to the working principle of the molecular pump. In addition, it should be noticed that the intensity of both Mo 3d and S 2p peaks varied with the measurement position actually. As there were a lot of MoS2 flakes on the tested SiO2 substrate, and tiny position shifts between adjacent measurements were inevitable, results obtained from different XPS measurements may contain information from different MoS2 flakes. Meanwhile, since the thickness and density of MoS2 flakes varied according to their locations, there were some intensity differences in both Mo 3d and S 2p components with treatment time. In this case, the intensity of Mo 3d and S 2p components with 5 + 15 s remote oxygen plasma treatment happened to be the maximum.1g modes for MoS2 at ~380 and 405 cm−1. According to the work of Bertrand P. A.1g mode is generated from the out-of-plane vibration of S atoms in opposite directions. From the inset of 1g modes can be observed before and after the sample undergoing the remote oxygen plasma treatments, implying that bonding situations of Mo and S atoms didn’t change. In addition, the peak that centers near 820 cm−1 could be used to estimate the extent of oxidation that occurred2 flakes were oxidized, this peak would be more defined and intense after the treatments. As expected, this peak showed no intensity difference before and after the remote O2 plasma pretreatments, indicating that the MoS2 flakes remained un-oxidized during the pretreatments. From these results, it is clear that during the treatments, the remote O2 plasma is gentle enough to avoid damaging the MoS2 flakes. Instead, the produced oxygen species are adsorbed onto the MoS2 surface and serve as nucleation sites for the initial TMA pulses during the ALD process.To further verify the results obtained by XPS, Raman spectra of the sample before and after the oxygen plasma treatments mentioned above were obtained in air using a 514-nm laser . The ins2-based field effect transistors were fabricated to estimate the impact of the remote O2 plasma pretreatment on the device mobility. A cross-sectional schematic of the MoS2 transistor with the remote O2 plasma treatment is shown in 2 flake is about 8.4 nm as shown in 2 channel by applying a source-drain bias (Vds) to the pair of electrodes as shown in bg) to the heavily doped silicon substrate. As shown in the insets in ds – Vds curves are all linear in the range from −40 to 40 mV with or without the remote O2 plasma pretreatment, indicating that the Cr/Au contacts are ohmic contacts. The transfer and output characteristics for the MoS2 transistor before and after 60 s remote O2 plasma pretreatment are obtained for comparison. The data presented in on/Ioff) over 107. This high on/off ratio compared to graphene transistors is attributed to the large band gap of MoS2. It is also observed in 2 plasma pretreatment. For example, the transfer current at Vds = 500 mV increases from 1.56 × 10−5 to 3.38 × 10−5 A after a 60 s remote O2 plasma pretreatment. A low field-effect mobility is extracted using the Equation:[1]i = 1.15 × 10−8 F/cm2 is the capacitance density between the channel and the back gate (details of the mobility extraction can be found in Supplementary Information). Results show that the mobility increases from the original value of 22.15 to 33.57 cm2/Vs after 60 s remote O2 plasma pretreatment. Moreover, from the comparison between 2 plasma pretreatment. Taking the Ids – Vds curve under Vbg = 40 V for example, the output current increases from 2.81 × 10−5 to 9.07 × 10−5 A after 60 s remote O2 plasma pretreatment. Furthermore, for both the transfer and the output characteristics, excellent field-effect behavior is observed.As the mobility is of great significance when evaluating the performance of electronic devices, back-gated MoSThe evolution of the device mobility is tested with different pretreatment time. As shown in 2 transistor with top gate dielectric deposited using remote oxygen plasma pretreatment. Top gate dielectric of this device is 60 cycles Al2O3 (about 6.6 nm) which was deposited at 300 °C with 60 s remote oxygen plasma pretreatment. L = 1 μm and W = 5 μm are the channel length and channel width, respectively. For all the measurements in ds and Vds within −40 mV −40 mV indicates that Cr/Au electrodes form perfect ohmic contacts with the MoS2 channel. In addition, for all the transfer curves presented in 8 can be observed within the ±3 V range of the top gate voltage. Top gate leakage current is also measured in the same device. Compared to previously reported top-gate leakage of 2 pA/μm2 within 2 MV/cm1−13 A (about 0.1 pA/μm2) in the measurement range of −3 V to 3 V (4.5 MV/cm). This leakage is much smaller and at same time with an ultrathin gate oxide. The field effect mobility of this top gate device is extracted using 2/Vs under Vds = 0.5 V with the SS to be 101 mV/dec.2O3 growth on the MoS2 basal plane was successfully achieved by applying a remote O2 plasma pretreatment before ALD, and the mechanism was investigated systematically. After a remote oxygen plasma pretreatment, the oxygen species are physically adsorbed onto the surfaces of the MoS2 flakes and act as nucleation sites for the ALD cycles. The transport studies reveal an extra benefit of this method, which is that unlike many other methods that might sacrifice the device mobility to achieve uniform high-κ growth, this method improves the device mobility by 50%. Furthermore, top-gated MoS2 transistor with ultrathin Al2O3 dielectric was also fabricated. With only 6.6 nm Al2O3 as dielectric, which is the thinnest top gate dielectric ever reported so far, the device shows impressive leakage about 0.1 pA/μm2 at 4.5 MV/cm. Besides, the top-gated device shows great on/off ratio of over 108, subthreshold swing (SS) of 101 mV/dec and a mobility of 28 cm2/Vs. According to the mechanism, it is believed that this method can also be adopted for high-κ growth on other two dimensional nanostructures and used in other devices. With further investigations and optimizations, this method could play an important role in the future nanoelectronics.In summary, uniform Al2 were obtained from bulk crystals (SPI supplies Brand) using the classical tape-based mechanical exfoliation method commonly used for graphene, then transferred onto degenerately doped Si substrates covered with 300 nm SiO2. The thicknesses of these flakes were determined with a Bruker Multimode 8 atomic force microscope (AFM).Ultrathin layers of MoS2 flakes were loaded into the Picosun R200 ALD chamber for direct Al2O3 deposition. During the deposition, TMA and H2O served as the aluminum and oxygen precursors, respectively, and different growth temperatures and pulse time were adopted to observe their impacts. For some of the flakes, the remote O2 plasma pretreatments were carried out in the same chamber before Al2O3 was deposited. Here, “remote” means that the plasma source is located remotely from the substrate stage, such that the substrate is not involved in the generation of the plasma. It is then carried to the sample surface by the carrier gasin-situ characterization was SPECS XR50 X-ray source. Considering that the signal intensity was not so strong due to the low density of MoS2 flakes on the substrate and the signal intensity might be further weakened by using an X-ray monochromator, we finally carried out the in-situ characterization using a non-monochromatic XPS source. In the in-situ XPS measurements, all the spectra were taken using a Mg Kα X-ray source (hν = 1253.6 eV). The working pressure in the ultra-high-vacuum (UHV) chamber for the data acquisition was maintained at the magnitude of 10−10 mbar. The element library and the quantification factors used during measurements were provided via the system SpecsLab2 software, and after the measurements, the data analysis was carried out using CasaXPS software. The binding energies in the XPS spectra were calibrated in the conventional way against the adventitious carbon C 1s singlet (Eb = 284.6 eV). The Raman spectra of the MoS2 flakes before the remote O2 plasma treatment were measured with a Renishaw inVia Raman microscope in air using a 514 nm laser. Then after going through the in-situ XPS characterization mentioned above, Raman spectra of the same sample was measured again with all the measurement settings to be the same.Some of the MoS2 layer served as the back gate and the gate dielectric, respectively. The source and drain contacts were formed using electron-beam lithography followed by deposition of 10 nm Cr and 70 nm Au. The electrical properties of the transistors were measured with an Agilent B1500 semiconductor device parameter analyzer.The degenerately doped Si substrate and the 300-nm SiOHow to cite this article: Yang, W. et al. The Integration of Sub-10 nm Gate Oxide on MoS2 with Ultra Low Leakage and Enhanced Mobility. Sci. Rep.5, 11921; doi: 10.1038/srep11921 (2015).


II6L4 capsules result from the subcomponent self‐assembly of 2‐formylphenanthroline, threefold‐symmetric triamines, and octahedral metal ions. Whereas neutral tetrahedral guests and most of the anions investigated were observed to bind within the central cavity, tetraphenylborate anions bound on the outside, with one phenyl ring pointing into the cavity. This binding configuration is promoted by the complementary arrangement of the phenyl rings of the intercalated guest between the phenanthroline units of the host. The peripherally bound, rapidly exchanging tetraphenylborate anions were found to template an otherwise inaccessible capsular structure in a manner usually associated with slow‐exchanging, centrally bound agents. Once formed, this cage was able to bind guests in its central cavity.Pseudo‐octahedral M Abiological molecules that can recognize, process, and transport chemical species provide insight into the fundamentals of complex natural chemical systems.Although most often designed to encapsulate guests centrally, a few cages have the ability to bind molecules in more than one mode or location.Templation, likewise, typically involves a series of subunits assembling around a central structure‐defining feature.II6L4 architectures were generated by employing 2‐formylphenanthroline (a tridentate component) in place of 2‐formylpyridine (a bidentate component) during the subcomponent self‐assembly process. Binding investigations of these hosts with a range of tetrahedral, octahedral, and icosahedral guests revealed that one cage could simultaneously bind two guests in two different locations, and that peripheral guests bound more strongly than internal ones.Herein, we present the syntheses and host–guest chemistry of a new class of supramolecular pseudo‐octahedra capable of binding neutral and anionic guests, in both internal and peripheral pockets. These new MII6L4 structure, despite the failure of this structure to form either directly by subcomponent self‐assembly or through the use of guest templation with centrally encapsulated guests. Once formed by peripheral templation, this capsule was capable of encapsulating guests centrally. This observation highlights the importance of peripheral templation in the generation of otherwise inaccessible host–guest species.In one case, peripheral guests were observed to template the formation of a Cd−; 6 equiv), and either triamine A or B in CH3CN led to the formation of ZnII6L4 assemblies 1 or 2, respectively, after heating for 16 h at 70 °C .The reaction of 2‐formylphenanthroline 12 equiv), zinc(II) trifluoromethanesulfonate . ESI‐MS confirmed the 4:6 metal/ligand ratio of the structure, while NMR spectroscopy revealed a highly symmetric species, as with 1 and 2. HR‐ESI‐MS unambiguously established the stoichiometry in all cases.Under all conditions tried, 1 and 2 were grown by slow diffusion of Et2O or iPr2O, respectively, into CH3CN solutions.II6L4 complexes comprise an octahedral framework of metal ions, in which the octahedron faces are alternately occupied by a ligand or an open aperture  point symmetry.Single crystals of 2 presents a more distorted octahedral ZnII coordination environment than in 1; the angles between the chelate planes of the imino‐phenanthroline units were observed to be 82–90° in 1, versus 71–90° in 2. Similarly, whereas 1 displayed uniform diagonal ZnII–ZnII distances of about 17 Å, the corresponding diagonals in 2 measured approximately 15×16×19 Å,2. Although of similar areas (20–30 Å2), the open triangular apertures of 2 resultantly present a less equilateral surface than 1. Void volumes, calculated using VOIDOO,3 for 1 and 423 Å3 for 2. These volumes are approximately ten times larger than those of the MII4L4 tetrahedra formed from the same triamines using 2‐formylpyridine.The crystal structure of 1–3 to bind tetrahedral guests. Three classes of tetrahedral prospective guests were investigated: neutral molecules, smaller anions, and larger anions incorporating aromatic units. Whereas the anions BF4−, ClO4−, SO42−, and PO43− were not observed by 1H or 19F NMR spectroscopy to bind within 1–3 at 25 °C, the addition of polyhalogenated CI4, CBr4, or CI3H led to shifts in the 1H NMR spectrum of 2, consistent with binding in fast exchange on the NMR timescale . As the most pronounced downfield shifts were observed for the phenylene protons of 2, we infer that binding occurs centrally. No binding was observed for the smaller CHBr3, CCl4, or less symmetric CH2BrI, CHCl3, or CH2Cl2 molecules  was observed to bind in intermediate exchange on the NMR timescale  with 1 and 3, whereas this anion was observed by 1H NMR spectroscopy to bind in fast exchange with 2 . Significant shielding of the phenylene protons of the ligand was observed in all cases, indicative of guest interaction at the face of the structure; however, substantial shifts for signals attributed to the imine and its nearest phenanthroline protons were also observed. In both 1 and 3, upfield shifts exceeding 2 ppm were observed, consistent with strong electronic perturbation at the windows of the architecture.Tetraphenylborate , or peripheral binding, where only one phenyl ring enters the cavity and the remaining three sit in pockets formed by the bisphenanthroline corners  indicated strong NOE correlations between the ortho protons of BPh4− and the phenylene, imine, and adjacent phenanthroline protons of 1–3. Irradiation of the meta BPh4− protons revealed considerably weaker NOE interactions with the phenylene protons of 1–3 and the two phenanthroline protons closest to the imine; no NOE signals were observed between the para BPh4− protons and 1–3. Consideration of the molecular models of BPh4−⋅1 revealed that only in the peripherally bound structure were the ortho protons of BPh4− sufficiently close to the phenanthroline corners to produce the observed NOE correlations . Although adjacent to the phenylene rings of the ligand, the ortho protons of an internally bound BPh4− would be too far away for NOE interactions to be observed to the imine or phenanthroline protons. Consequently, a peripheral binding model, in which a phenyl ring of the guest protrudes through a window of the architecture, best describes the binding of BPh4− ions to 1–3. This mode is distinct from the external binding described by Raymond and co‐workers, wherein guests undergo nonspecific interactions with the exterior of the cage and are not directly bound in a cavity.Tandem 1D selective 1–3, we treated these hosts with tetra‐p‐F‐ and tetra‐p‐Cl‐substituted tetraphenylborate anions (B(C6H4F)4− and B(C6H4Cl)4−), both of which were observed to bind in fast exchange on the NMR timescale to 1–3 . No binding was observed for pentafluoro‐ or bis‐m‐CF3‐substituted tetraphenylborates, or for the structurally analogous tetraphenylmethane 4−⋅1  furthermore revealed NOE interactions between the para‐fluorine substituent of the guest and the protons H5 and H6 of the phenanthroline moiety. These NOE interactions are consistent with the phenyl rings of B(C6H4F)4− resting on the triangular corners of 1. No such NOE interactions are possible should the guest be internally bound.A 1 and 3, and 1H NMR spectroscopy for 2) were fitted using 1:1 binding isotherms . Monitoring the titration of BPh4− into 3 by cyclic voltammetry  resulted in a positive shifting of the first and second reduction waves, attributed to the reduction of the imino‐phenanthroline motif. These observations are consistent with a donation of electron density from the guest to the host, and a narrowing of the band gap upon anion binding.Although 2; downfield shifting of its phenylene protons, along with movement of the imine and adjacent phenanthroline protons, indicated proximity of the anions to the central cavity of the cage . Broadening of the 19F signals for AsF6− and SbF6− was likewise observed, consistent with binding being in fast exchange on the NMR timescale. These anions exhibited only weak association, however, with binding affinities <40 m−1, as determined through 1H NMR titrations , which bound in fast exchange with 2 on the NMR timescale . The B12H122− and B12F122− dianions were also observed to bind within 2 in fast exchange by 1H NMR spectroscopy , although their limited solubilities precluded the quantification of their binding affinities. Nevertheless, splitting and downfield shifting of the overlapping signals corresponding to the phenylene protons of 2 upon titration with B12F122− and CB11H12− ions indicated their uptake within the cage cavity. Treatment of 1 or 3 with the same octahedral and icosahedral anions led either to no significant change in their 1H NMR spectra or to broadening, which could not be resolved even at −40 °C.A higher affinity for 2 possesses multiple binding sites that each bind specific anionic guests, we explored the allosteric effects engendered by treating 2 simultaneously with a guest specific to each site (see Section 10 in the Supporting Information). The titration of CsCB11H12 into a solution of BPh4−⋅2 in CD3CN did not lead to a significant shift in the 1H NMR signals of the bound BPh4− (Δorthoδ=−0.02 ppm), thus suggesting that concurrent binding of exo‐BPh4− and endo‐CB11H12− had occurred. Furthermore, the presence of peripherally bound BPh4− had no significant effect on the binding affinity of CB11H12− to the inside of 2 (Ka=1.59±0.08×102 m−1), nor did the presence of endo‐bound CB11H12− considerably alter the binding strength of BPh4− to the periphery of the cage (Ka=2.8±0.4×103 m−1). Increasing the concentration of the other guest in both cases did not change the binding constant of either the endo‐ or exo‐bound guest.Having thus established that host 12F122− and BPh4− were likewise observed to bind to 2 simultaneously. The addition of K2B12F12 to a solution of BPh4−⋅2 led to 1H and 19F NMR shifts consistent with the encapsulation of B12F122−. Reversing the order of titration by adding nBu4NBPh4 to B12F122−⊂2 did not significantly alter the 19F NMR chemical shift of encapsulated B12F122−, thereby indicating that B12F122− was not ejected from the capsule upon the peripheral binding of BPh4−. The system thus appears to bind internal and peripheral guests concurrently, with neither allosteric inhibition nor enhancement of binding affinity.B1–3 proved ineffective for the synthesis of the marginally larger structure 4. Heating 2‐formylphenanthroline (12 equiv), Cd(OTf)2 (6 equiv), and B (4 equiv) in CH3CN to 60 °C overnight resulted in a broad, ill‐defined aromatic region in the 1H NMR spectrum; only free B could be positively identified. Having elucidated the unique binding abilities of 2, we sought to use this knowledge to generate 4 through guest templation. Although the addition of internally binding guests resulted in no significant change in the broad 1H NMR spectrum of the precursors of 4, the addition of nBu4NBPh4 (2 equiv) led to the development of signals corresponding to the host–guest species BPh4−⋅4 during 12 h of heating at 50 °C . The diffusion coefficient of capsule 4 was measured to be 9.2×10−8 m2 s−1 by DOSY NMR spectroscopy , similar to the values found for 1 and 2; BPh4− bound to 4 was observed to diffuse more slowly than free BPh4−. HR‐ESI‐MS confirmed the expected MII6L4 stoichiometry of the resulting species . Notably, no MS signals corresponding to the free cage (without BPh4−) could be identified, which reflects the strong binding of this anion to 4.Similar conditions to those used for the syntheses of para‐substituted tetraphenylborate templates likewise generated 4, although two or more equivalents of template were consistently necessary for the formation of the host in all cases . Templation did not, however, occur in the presence of any centrally binding guest; the addition of CB11H12− or B12F122− to its precursors did not produce 4.The use of 4−⋅4 revealed the same mode of peripheral binding as was observed in the cases of 1–3 . The ability of BPh4− to template the formation of 4 is thus highly unusual—a template typically sits at the center of the chemical structure that it brings into being, forming a symmetrical host–guest adduct. Furthermore, the high symmetry reflected in the 1H NMR spectra of the adducts of 4 with tetraphenylborate and its halogenated congeners indicates that the guest remains in fast exchange on the NMR timescale between different sites on the host at 25 °C. We are not aware of another example of a rapidly exchanging, exo‐bound agent templating the formation of a metal–organic capsule.NOE measurements on BPh4, once formed, 4 was observed to bind CB11H12−. The titration of CsCB11H12 into a solution of BPh4−⋅4 in CD3CN resulted in 1H NMR shifts consistent with the encapsulation of the carborane anion, with a simultaneous shift in the protons of BPh4− . The binding affinity of CB11H12− for BPh4−⋅4 was calculated to be 1.37±0.03×102 m−1, similar to its ZnII analogue 2.Although carborate did not serve as a competent template for the formation of 4, which could not be achieved in the template's absence. Moreover, the inclusion complex BPh4−⋅4 could still participate in host–guest chemistry. This new mode of templation, which is allosteric in nature, enables the exploration of more complex chemical systems that are capable of responding to multiple stimuli.Employing 2‐formylphenanthroline in place of 2‐formylpyridine during subcomponent self‐assembly thus led to the formation of larger structures with significantly expanded cavities. Understanding the host–guest chemistry of these structures enabled the development of a new mode of templation. The addition of a rapidly exchanging, peripherally bound guest engendered the formation of As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re‐organized for online delivery, but are not copy‐edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.SupplementaryClick here for additional data file.


M) and vehicle‐(DMSO 0.05%) treated hippocampal slices obtained from mice. The soluble Aβ treatment did not produce alterations in sub‐threshold intrinsic properties, including membrane potential, input resistance, and hyperpolarization activated “sag”. Similarly, no changes were noted in the firing profile evoked by 500 ms square current supra‐threshold stimuli. However, Aβ 500 nM treatment resulted in the hyperpolarization of the action potential (AP) threshold. In addition, treatment with Aβ at 500 nM depressed the after‐hyperpolarization that followed both a single AP or 50 Hz trains of a number of APs between 5 and 25. These data suggest that acute exposure to soluble Aβ oligomers affects IE properties of CA1 pyramidal neurons differently from outcomes seen in transgenic models of amyloidopathy. However, in both chronic and acute models, the IE changes are toward hyperexcitability, reinforcing the idea that amyloidopathy and increased incidence in seizures might be causally related in AD patients. © 2014 The Authors Hippocampus Published by Wiley Periodicals, Inc.Accumulation of beta‐amyloid (Aβ) peptides in the human brain is a canonical pathological hallmark of Alzheimer's disease (AD). Recent work in Aβ‐overexpressing transgenic mice indicates that increased brain Aβ levels can be associated with aberrant epileptiform activity. In line with this, such mice can also exhibit altered intrinsic excitability (IE) of cortical and hippocampal neurons: these observations may relate to the increased prevalence of seizures in AD patients. In this study, we examined what changes in IE are produced in hippocampal CA1 pyramidal cells after 2–5 h treatment with an oligomeric preparation of synthetic human Aβ 1–42 peptide. Whole cell current clamp recordings were compared between Aβ‐(500 n Beta‐amyloid peptides (Aβ) are widely believed to play an important role in the pathogenesis of Alzheimer's disease (AD)  and to date it has involved investigation of four different transgenic Aβ overproducing mouse lines. We have identified changes to the bursting behavior of CA1‐PC and associated changes to the spike after‐depolarization (ADP) . The concentration used in this study has been frequently used for in vitro studies of synaptic function and it has been shown to both impair : sucrose, 189; D‐glucose, 10; NaHCO3, 26; KCl, 3; MgSO4, 5; CaCl2, 0.1; NaH2PO4, 1.25, continuously bubbled with carbogen . The cerebellum, frontal and dorsal parts were removed with single scalpel cuts. The sample was then mounted on a metal plate on the dorsal side  and 300 μm thickness horizontal sections were prepared using a Leica VT1200 vibratome. After sectioning, slices were submerged in a storage vessel, which contained our standard artificial cerebrospinal fluid (aCSF) consisting of (in mM): NaCl, 124; KCl, 3; NaHCO3, 26; CaCl2, 2; NaH2PO4, 1.25; MgSO4, 1; d‐glucose, 10, and equilibrated with carbogen. The slices were gradually heated to ∼32–34°C for 30 min, after which they were stored at room temperature for 1 h before being treated with either hAβ 1–42 peptide or vehicle.All procedures were performed in accordance with UK Home Office legislation set out in the Animals (Scientific Procedures) Act (1986). The preparation of horizontal ventral hippocampal slices was performed as previously described  for 1 h, vortexing every 5–10 min. The preparation was then sonicated for 10 min at RT. The HFIP/peptide solution was subsequently dried down under a gentle stream of nitrogen. Following this, the dried peptide was re‐suspended in 100% DMSO to a concentration of 1 mM and incubated for 12 min at RT, gently rotating the vial and vortexing every few minutes. This preparation was then pipetted into 10 μL aliquots and stored at −80oC. On the day of use, a single aliquot was rapidly thawed and 90 μL of PBS 0.1M added before performing 90–120 min of rotation and gentle agitation. Following this, the Aβ preparation was finally diluted 1:2000 into standard aCSF to a final Aβ concentration of 500 nM and applied to slices for 2–5 h before electrophysiological recording. Control slices were exposed for a similar period to vehicle only (0.05% DMSO); use of this vehicle control group is particularly important as we have shown that this concentration of DMSO can alone change IE of cortical and hippocampal neurons  with carbogen‐equilibrated aCSF at 33 ± 1°C. CA1 pyramidal neurones in the CA1 subfield of the hippocampus were visually identified using infra‐red (IR) differential interference contrast optics. Pipettes were fabricated from borosilicate glass and were fire polished such that their resistance was 2.5–4.5 MΩ when filled with pipette solution. The pipette solution consisted of (mM): K‐gluconate, 135; NaCl, 5; HEPES free acid, 10; EGTA, 0.2; Na‐GTP, 0.3; Mg‐ATP, 4; Biocytin 13.4, Alexafluor 488 hydrazide 0.03–0.06, pH 7.3, 280–290 mOsm.Effects of exogenous Aβ on IE were studied using single cell patch clamp recording from CA1‐PC. The recording methods we used are similar to those we used for our previous studies of intrinsic properties of this cell type in both Aβ‐overproducing transgenic mice and healthy aged animals . Recordings were low pass filtered (5–10 kHz) and then digitised (20–100 kHz) with a Digidata 1440 and stored on a personal computer using pClamp10 electrophysiology software.Rin‐exp) assessed resting input resistance independently of the “sag”‐producing Ih activation that occurs during hyperpolarizing current steps. This was calculated using Ohm's law from the amplitude of an infinite time extrapolation of a single exponential curve fitted to the membrane charging response generated by a −100 pA current injection  was calculated using Ohm's law by determining the steady‐state (post‐sag) voltage deflection produced by a 500 ms, −100 pA stimulus  was also measured  using linear regression of the steady state voltage responses elicited by a series of eight low amplitude (−50 to +30 pA), 500 ms duration current steps  using the appropriate amount of current injection  simply expressed the difference between the peak  measured the decay in response relative to the amplitude of the infinite time extrapolation used to determine Rin‐exp i.e., 100* (B–C)/B /I(fft); the impedance versus frequency profile was subsequently smoothed with a moving average function with a span of 35 data‐points. The parameters analyzed to test the effect of treatment on the sub‐threshold resonance properties of CA1 pyramidal neurons were the peak frequency, the peak impedance and the quality factor of the resonator Q; unpaired Student's t‐test was used to test the effect of treatment on resonance.Membrane resonance was measured with standard “ZAP” protocols, in which a low amplitude sinusoidal current injection of linearly increasing frequency was applied to the cell causing voltage deflections of around 5–10 mV, as previously described  firing. From these, the relationship between the stimulus amplitude and the number and pattern of APs elicited was examined. The firing properties illustrated in Figures t‐test was used to compare the ADP and AHP elicited by a single spike.Voltage responses were also measured to single, brief (2 ms), strong (2 nA) current injections. This stimulus usually elicits a single AP, which is followed in hippocampal pyramidal cells by a fast ADP, followed in turn by a brief small after‐hyperpolarization (AHP). Unpaired Student's Finally, to further test the effect of treatment on post‐burst firing AHP, trains of 5–10–15–20–25 2 ms/2 nA square current pulses were delivered at a frequency of 50 Hz; the medium AHP was measured as the most hyperpolarized point in the 500 ms following the descending phase of the last AP reaching 0 mV. Two way ANOVA was used to statistically assess the effect of treatment on the medium AHP amplitude elicited by 50 Hz spike trains.M peptide . Although high compared to global average levels in AD brain measured in Aβ‐overexpressing transgenic mouse models of amyloidopathy (∼0.5 nM)  relies on the fact that the average levels of Aβ 1–42 soluble oligomers in the AD brain can disguise the peri‐plaque concentration gradient of this species, which might differentially affect the neuronal function depending on the distance from the plaque itself.Our studies of the effects of exogenous Aβ on IE of CA1‐PC were focused on slices treated with 500 n Waters, , this co Waters, . The con Waters, . The phyM of Aβ had no effect on resting membrane potential  was unaltered  concentration of hAβ 1–42 oligomers, was not able to alter the AP by AP profile of instantaneous frequency in a way similar to that we reported in the Aβ overproducing transgenic mice . Thus, an acute 2–5 h treatment with 500 nM hAβ 1–42 oligomers was not sufficient to produce the same changes in IE and AP firing previously observed in Aβ‐overproducing transgenic mice.The post‐spike ADP is a major determinant of burstiness in hippocampal pyramidal cells, and was larger in PSAPP mice  treatment (n = 9) compared to vehicle treated slices . Similarly the AHP observed in CA1‐PC after a single AP (elicited with a single 2 nA current injection lasting 2 ms) was also around 50% smaller in Aβ treated slices (control: 1.2 ± 0.2 mV (n = 17) versus Aβ: 0.6 ± 0.1 mV, (n = 17), P < 0.02).To investigate how Aβ affected AHPs following bursts of AP firing we used repeated brief, strong current injections  to induce CA1‐PCs to fire trains of AP at 50 Hz. The trains consisted of 5, 10, 15, 20, or 25 APs. In CA1‐PC from control slices such stimuli reliably evoked a mAHP; in contrast, mAHPs were completely absent in many cells from Aβ treated slices. Furthermore, in the peptide treated cells a postburst ADP not seen in control cells and lasting a few hundred ms was apparent. This is illustrated in Figure in vivo  or vehicle alone . It appears especially important to perform the correct vehicle control in studies such as this as we have recently demonstrated that 2–5 h treatment with 0.05% DMSO alters IE properties of both CA1‐PC and pyramidal neurons in perirhinal cortex  left many basic intrinsic properties of CA1‐PC unaltered, including resting potential, input resistance and any other sub‐threshold property. Aβ treatment did, however, produce both a negative shift in action potential threshold and a depression of the substantial mAHP that usually arises after 50 Hz trains of AP firing. Indeed, in many cells the mAHP was not only lost but appeared to be replaced with a slow ADP.In previous reports by other groups, certain neurophysiological effects of Aβ peptides have been reported to be detectable within short exposure periods. For example, the inhibition of long‐term potentiation is reported to require less than 1 h of Aβ pre‐treatment and maybe as little as 10 min A more hyperpolarized AP threshold and a depressed AHP (and appearance of an ADP) after spike firing are both actions that would typically be regarded as a net increase in excitability. For the former this is because following Aβ exposure a smaller change from resting potential would be required to induce AP production. For the latter, the changes to spike afterpotential would enable spiking to be re‐established more readily after a prior burst. Certainly, hyperexcitability in cortical and hippocampal circuits of Aβ over‐expressing mice is an increasingly reported phenomenon, and has been discussed in the framework of the increased risk of seizures found in AD suffers, most notably in early onset cases: the changes to intrinsic properties described here may contribute to this phenomenon (Brown et al., An interesting insight from these results is the comparison with transgenic models of amyloidopathy, where the exposure of CA1 pyramidal neurons to increased Aβ is chronic, typically lasting months or longer. Recent studies showed that PSAPP (Brown et al., M Aβ 1–42 concentration used in this study for acute slice pre‐treatment. Indeed, in terms of clinical AD, the commonly used Aβ concentration we used to treat slices with is high. Mean global levels of soluble Aβ 1–42 in the brains or CSF of AD sufferers are typically in the low pM range. It remains possible, however, that higher levels are present within certain CNS niches, for example in the vicinity of amyloid plaques or perhaps within synaptic clefts.The global Aβ concentration in the brain of Aβ overproducing transgenic mice is likely to be much lower than the 500 nWhen presenting our previous research on transgenic mice that overproduce Aβ we have often been asked if we observe similar behaviors when applying soluble Aβ peptides acutely. By performing many similar measurements to those we made previously in PSAPP (Brown et al., + channels. This is something which could possibly be investigated with voltage‐clamp studies of nucleated macropatches, an approach we have used previously to identify Na+ channel changes underlying altered IE both in mouse models of amyloidopathy (Brown et al., 2+ in CA1‐PC AHPs, the altered afterpotentials seen following Aβ treatment may reflect a change to activity‐dependent intracellular Ca2+ signalling, something that might best be examined further with cellular imaging methodologies.It may be useful in future to investigate the cellular basis of the changes to IE that are produced by acute Aβ treatment. The altered AP threshold is likely to be a consequence of a change to voltage‐gated Na


We find it to be basically kinetic and highly complex, involving non-negligible contributions up to seventh power of total angular momentum of each lanthanide site. The performed analysis also elucidates the origin of magnetization blocking in these compounds. Contrary to general expectations the latter is not always favored by strong exchange interaction.Combining strong magnetic anisotropy with strong exchange interaction is a long standing goal in the design of quantum magnets. The lanthanide complexes, while exhibiting a very strong ionic anisotropy, usually display a weak exchange coupling, amounting to only a few wavenumbers. Recently, an isostructural series of mixed  For strongly axial doublet states on the lanthanide sites (Ln) these Hamiltonians basically become of Ising typeThe key feature of lanthanide ions in materials is their strong magnetic anisotropy caused by strong spin-orbit coupling effectsz1 ‖ z2) or non-collinear either collinear (3Si)2N] (THF)Ln}2 (μ − η2:η2 − N2) (Ln = Gd (1), Tb (2), Dy (3), Ho (4), Er (5), THF = tetrahydrofuran), shown in Fig. 1a14J-multiplets on the lanthanide sites, implying that the picture of exchange interaction involving individual crystal-field doublets, This paradigm was recently challenged by a series of ab initio and density-functional theory (DFT) calculations with microscopic model description to unravel the nature of this exchange interaction. We also elucidate the origin of blocking barriers in these compounds and discuss the effect of strength of exchange interaction on magnetization blocking in strongly anisotropic complexes.The purpose of the present work is to reveal the mechanism of giant exchange interaction and the origin of the magnetization blocking of the series of the complexes based on adequate theoretical treatment. We apply an approach combining 3+ ions (SGd = 7/2) interact with the radical spin of the To understand the origin of such strong exchange interaction, we consider the simplest complex of the series, the gadolinium one. In this system the isotropic spins of Gd complex . Broken-Dh2 symmetry of the exchange core  to both lanthanides. Most important, this orbital is found to lie higher than the 4fxyz orbitals by as much as Δ = 5.2 × 104 cm−1 7 (π*)1 → (4f)8 (π*)0 → (4f)7 (π*)1, the kinetic contribution to the Gd3+-To get insight into the mechanism responsible for the obtained huge value of nge core , the ant xyz one . The cor104 cm−1 . Becaused orbitals of Gd3+  order of the perturbation theory compared to the usual kinetic exchange, and involves the excitation energy into a higher 5d orbital on the Ln site. Both these contributions are usually neglected unless the conventional kinetic exchange appears to be small293+-−1.Compared to this mechanism, the other contributions such as the direct exchange, the delocalization of unpaired electron of f and the π* orbital levels is obtained as huge as in the Gd complex  is of the same order of magnitude as the estimated isotropic exchange splitting in 1 (≈400 cm−1). Therefore, in sharp contrast with the common situation in lanthanides, the exchange coupling in the anisotropic 2–5 does not reduce to the interaction between individual (lowest) CF doublets on Ln sites with the S = 1/2 spin of the radical, J multiplet at lanthanide ions. Then, such exchange interaction should be formulated in terms of the total angular momenta i = 1, 2) on the lanthanide sites.Contrary to the Gd complex, the other members of the series are characterized by strong magnetic anisotropy on the Ln sites induced by the crystal-field (CF) splitting of their atomic 3+-3+-3+ sites Extending the Anderson’s superexchange theory29S = 1/2, k and component q, and α = x, y, z) of kth degree, in which |q| (=±q) corresponds to the order of k is 7 for the considered Ln3+ ions, whereas the maximal |q| is 5 in the present case because only the 4fxyz magnetic orbitals at the lanthanide sites contribute to the kinetic exchange. The summation over k in J multiplets on individual metal sites, whereas S = 1/2 of the D is the 3 × 3 exchange matrix, containing one isotropic, five symmetric anisotropic and three antisymmetric  exchange parametersk = 1) in D. The expression for the exchange parameter f)n (π*)1 → (4f)n+1 (π*)0 → (4f)n (π*)1, where n is the number of 4f electrons in Ln3+. The multiplet electronic structure of Ln2+ is fully included in the electron promotion energy U0 + ΔEα and the wave functions of the intermediate states, where by U0 we further denote the smallest promotion energy, α numbers the intermediate J-multiplets, and ΔEα is the excitation energy of the multiplet α with respect to the ground one in Ln2+.Here, t is obtained here from DFT calculations, expected to be accurate enough39Eα and the CF states are obtained by fragment state-of-the-art ab initio calculations including spin-orbit coupling32ab initio calculations is U0. Indeed, the former gives at most an averaged value over multiplets The highly complex tensorial form of the exchange Hamiltonian is inevitable for orbitally degenerate systems with strong spin-orbit coupling, as was pointed out long time ago37U0, where ab initio CF Hamiltonian for mononuclear Ln fragments  in the ground exchange KD, the squares of which characterize the rate of quantum tunneling of magnetization (QTM)4 and the smallest for 2 and 3 complexes. This explains why large magnetization hysteresis is seen at low temperatures in the latter two compounds, while not seen at all in the former and only weakly observed in the complex 5 14Ebarrier corresponds to the first excited exchange KD, because its two components  approach was used including all seven 4f orbitals of the Ln atom in the active space. The spin-orbit interaction was treated with the module SO-RASSI and the local magnetic properties were calculated with the SINGLE_ANISO module of Molcas33Energies and wave functions of CF multiplets on LnFor further details, see How to cite this article: Vieru, V. et al. Giant exchange interaction in mixed lanthanides. Sci. Rep.6, 24046; doi: 10.1038/srep24046 (2016).


This article was initially published in the Epidemiology and Health 2016;38:e2016010, with an error in Figure 1C that duplicated Figure 1E.The authors would like to correct Figure 1C as below.


Optical properties of Ge5As37S58, Se layers, and Ge5As37S58–Se multilayer nanostructures were investigated. Spectral dependencies of refractive index were analyzed within the frames of single oscillator model. Values of optical band gaps for Ge5As37S58, Se layers, and Ge5As37S58–Se multilayer nanostructures were obtained from Tauc dependencies. Using e-beam and holographic recording, diffraction gratings were fabricated in Ge5As37S58–Se multilayer nanostructures. Images of Ukraine and Moldova state emblems were obtained by e-beam recording. Image size consisted of 512 × 512 pixels (size of 1 pixel was ~2 μm). Ge5As37S58–Se multilayer nanostructures are perspective for the direct recording of holographic diffraction gratings and other optical elements.Processes of e-beam and holographic recording of surface relief structures using Ge Thin films based on chalcogenide glasses have evolved as light-sensitive materials for high-density recording media application due to their optical and structural properties. The light sensitivity effect of thin chalcogenide films was discovered 50 years ago . Chalcog5As37S58–Se nanomultilayer structures under e-beam or holographic exposure are presented.In this work, the experimental results showing the surface relief formation in GeSelective etching after exposure enables to obtain surface reliefs and to use media such as high-resolution inorganic resists –7. UsingMultilayer nanostructures on the base of chalcogenide glasses and the possibility to use them as registering media were proposed in . Such me5As37S58–Se multilayer nanostructures for the fabrication of surface reliefs [5As37S58–Se multilayer nanostructures as recording media are presented.Chalcogenide glasses of Ge–As–S composition are characterized by high values of refractive index, and their nonlinear optical properties are two orders higher than characteristic of quartz glasses , 18. Ear reliefs . In pres5As37S58 were fabricated by common melt quenching method [5As37S58–Se nanomultilayers were prepared by computer-driven cyclic thermal vacuum deposition from two isolated boats with Ge5As37S58 and Se on constantly rotated glass substrate with deposited ITO layer at room temperature in one vacuum deposition cycle  of multilayer nanostructure. Overlapping part of the samples , and at the lowest photon energies [α ≥ 104 cm−1) which corresponds to transitions between extended states in both valence and conduction bands where the power law of Tauc [For chalcogenide semiconductors, the optical absorption coefficient, energies . The fir of Tauc  is validB is the slope of the Tauc edge which reflects some disorder of the samples. Usually, this constant depends on the width of the localized states in the band gap, a fact explained with the homopolar bonds’ presence in the chalcogenide glasses. Thus, Tauc plots of (αhν)1/2 versus (hν) should be linear and extrapolate to values of the optical gap, Eg.hv) is the photon energy, E0 is the oscillator energy, and Ed is the oscillator strength or dispersion energy. The parameter Ed which is a measure of the intensity of the inter-band optical transition does not depend significantly on the band gap. By plotting (n2 − 1)−1 against (hν)2 and fitting a straight line to the points, Ed and E0 can be directly determined from the slope, (EdE0)−1, and the intercept E0/Ed on the vertical axis. The extrapolation for (hv)→0 also gives frequency-independent refractive index or the so-called static refractive index, n0.The dispersion of the refractive index was analyzed using the Wemple–DiDomenico (WDD) model , 28, whiNc is the effective coordination number of the cation nearest neighbor to the anion, Za is the formal chemical valency of the anion, Ne is the total number of valence electrons per anion, and β is a constant . As found by Wemple and DiDomeniko [E0 is closely related to the optical band gap energy Eg.The dispersion energy is related to other physical parameters of material through the empirical formula:Domeniko , the osc5As37S58–Se multilayer nanostructures practically coincides with the absorption edge of Se  and the presence of scattering in Se layer .As can be seen from Fig. 5As37S58 and Se nanolayers were 16 and 14 nm, respectively, and are sufficiently smaller than the light wavelength. Also, transmission curve of Ge5As37S58–Se multilayer nanostructure is a typical interference curve for films with high optical quality and uniform thickness  were obtained.Obtained spectral dependencies of refractive index of Geαhν = const(hν − Eg)2, where hν is the light quantum energy, α absorption coefficient, and static refractive index, n0, are presented in Table Parameters of single oscillator model dispersion energy and effective oscillator position and also values of optical band gap obtained with the use of Tauc dependence 5As37S58–Se multilayer nanostructure are close. The Wemple–DiDomenico model in the range of the low-frequency optical dielectric response of glasses can be used as a tool to probe the building blocks conforming a glass and allows quantitative analysis in combination with Raman spectroscopy [It is necessary to note that values of optical band gaps  interdiffusion processes in nanolayers . The pro−1, obtained on the base of Ge5As37S58–Se multilayer nanostructure, is shown.In Fig. 2S3) are not additive in respect to thickness increase in the same conditions for Se–As2S3 multilayer nanostructure. The sum of the thickness increase in separate layers consists of only 30 % from the total measured thickness increase in Se–As2S3 multilayer nanostructures. In multilayer nanostructures, it is necessary to take into account also the presence of other processes, for example, formation of As–S–Se. It is supposed that stimulated by light or e-beams processes activate the mentioned processes in such media.Under exposure by ion- or e-beams of thin layers of chalcogenide, vitreous semiconductors structural transformations in films are observed –5, 36–38In Fig. 5As37S58–Se multilayer nanostructure are shown. Image sizes of the Ukraine state emblem consisted of 512 × 512 pixels (pixel size ~2 μm). AFM image of the recorded emblem fragment is shown on the right side of Fig. 5As37S58–Se multilayer nanostructure. AFM image of the recorded emblem fragment is shown on the right side of Fig. In Fig. 5As37S58–Se multilayer nanostructures, it is possible to realize direct recording of holographic optical elements (diffraction gratings) and also direct recording by e-beam exposure of diffraction gratings and other surface relief structures.Obtained results show that with the use of Ge


S. cerevisiae. In this study we examine the structural effects of a conserved serine residue in the P-loop corresponding to G12 in Ras.Sec4p is a small monomeric Ras-related GTP-binding protein (23 kDa) that regulates polarized exocytosis in We show that the Sec4p residue serine 29 forms a hydrogen bond with the nucleotide. Mutations of this residue have a different impact than equivalent mutations in Ras and can form stable associations with the exchange factor allowing us to elucidate the structure of a complex of Sec4p bound to the exchange factor Sec2p representing an early stage of the exchange reaction.Our structural investigation of the Sec4p-Sec2p complex reveals the role of the Sec2p coiled-coil domain in facilitating the fast kinetics of the exchange reaction. For Ras-family GTPases, single point mutations that impact the signaling state of the molecule have been well described however less structural information is available for equivalent mutations in the case of Rab proteins. Understanding the structural properties of mutants such as the one described here, provides useful insights into unique aspects of Rab GTPase function. S. cerevisiae and acts on the surface of membranes to regulate transport between the Golgi apparatus and the plasma membrane [Rab GTPases comprise the largest family of proteins among the small Ras-like GTPase superfamily functioning as critical regulators of a variety of membrane transport processes . Sec4p imembrane –4. The lmembrane –8. Membemembrane , 9–12. Mmembrane . As a remembrane , 15.The residue equivalent to RasG12 in Rab proteins is a serine that is well conserved throughout the Rab GTPase family . Cells were cultured at 37 °C until reaching an OD600nm of 0.6 followed by induction of protein expression using 50 μM IPTG and incubation at 15 °C for 14 h. Cells were harvested at 4000 × g for 30 min and resuspended in a lysis buffer containing 20 mM Tris-HCl pH 8.0, 300 mM NaCl, 5 mM MgCl2. The cells were lysed with 5 cycles of sonication while chilled in an ice bath. Unbroken cells and debris were pelleted at 18,000 × g for 30 min at 4 °C and supernatant solution was used for protein purification. The first round of purification was achieved by incubating the lysate with cobalt-conjugated beads (GoldBio Technologies). The protein was eluted in presence of buffer containing 20 mM Tris-HCl pH 8.0, 300 mM NaCl, 5 mM MgCl2 and 300 mM imidazole. The sumo tag was cleaved by adding SUMO protease to the eluted sample and the protein mixture was dialyzed against lysis buffer for 12 h at 4 °C. The digested sample was incubated with cobalt-conjugated beads and the unbound fraction containing the protein of interest was recovered. As a final purification step, the protein sample was concentrated and applied to a size exclusion column.The proteins used in this study were NHed Table . All DNA19 − 187S29V and Sec2p51–142 were first purified separately and then a 1.8-fold molar excess of Sec2p51–142 was mixed with Sec4p19 − 187S29V in buffer containing 20 mM Tris-HCl pH8.0 100 NaCl, 5 mM MgCl2 for 2 h at 4 °C. After incubation, the protein mixture was concentrated and applied into a size exclusion column Superdex200 (10/300)  to isolate the protein complex. Complex formation was evaluated by SDS-PAGE.Sec4p19 − 187S29V were performed at a protein concentration of 20 mg/ml. After initial crystal screening small crystals were obtained in presence of 14 % PEG3350 and 200 mM Zinc Acetate. After refinement of the initial condition, large single crystals were obtained in 14 % PEG4000, 50 mM Zinc Acetate with the addition of 10 mM GDP to the drop. Crystals were grown by the hanging-drop vapor-diffusion method after incubation at 20 °C for 10 days. Crystals were soaked in a cryoprotectant solution containing the mother liquor  supplemented with 20 % glycerol prior to flash-cooling in 100 K nitrogen gas stream. Diffraction data were collected in a rotary anode with crystal diffracting up to 1.9 Å resolution. The crystal belongs to space group P21, and Matthews’s coefficient calculation [Crystallization trials for Sec4p19 − 187S29V.GDP.Sec2p51–142 were performed at a protein concentration of 15 mg/ml. Crystals were grown by mixing protein complex with a reservoir solution containing 24 % PEG3350 0.2 M Sodium Citrate in a 1:1 ratio and with the addition of 10 mM GDP to the drop. Crystals were soaked in a cryoprotectant solution containing the mother liquor  supplemented with 20 % glycerol prior to flash-cooling in 100 K nitrogen gas stream. The crystallographic data was collected at the Brookhaven National Laboratories (BNL) National Synchrotron Light Source (NSLS). The crystal diffracted up to 2.9 Å resolution and belong to space group I222 with one ternary complex per asymmetric unit (% solvent).Crystallization trials for the ternary complex Sec4p19 − 187S29V.GDP.Sec2p51–142 was also solved with Phaser [All the data sets were processed using Mosflm  and Scalh Phaser  using a h Phaser . The strh Phaser , Phenix h Phaser  and manuh Phaser .2) with a 2-fold excess of mantGDP over the protein concentration and 5 mM EDTA. The mixture was incubated at room temperature for 10 min and the reaction stopped with the addition of 10 mM MgCl2. The unbound nucleotide was separated from the sample using a spin column  and the protein eluted in buffer containing 20 mM Tris-HCl pH8.0, 100 mM NaCl, 5 mM MgCl2. The protein concentration was measured by Bradford [In order to perform the nucleotide exchange experiments all the GTPases used in this work were preloaded with mantGDP. The procedure consists of incubating the protein sample . The mantGDP was excited at 360 nm and emission detected at 450 nm. For the intrinsic nucleotide exchange assay, time-based fluorescence was performed by collecting the fluorescence intensity in intervals specified in the plots. The data were normalized and plotted using Kaleidograph software (Synergy Software).Fluorescence measurements were performed in buffer containing 20 mM Tris–HCl pH8.0 100 mM NaCl, 5 mM MgCl2 at room temperature. The experiments were carried out using a stop flow apparatus coupled to a QuantaMaster™ 40 fluorescence spectrometer (PTI). The mantGDP was excited at 360 nm and emission was detected at 450 nm. The data were normalized and plotted using Kaleidograph software (Synergy Software).The fast kinetic experiments were performed in buffer containing 20 mM Tris–HCl pH8.0 100 mM NaCl, 5 mM MgClURA3 SEC2 plasmid. Yeast expressing wild type SEC2 can survive equivalently on the 5-FOA containing media, while a control plasmid with no insert cannot.In order to assess if various Sec2 constructs could act as the only copy of Sec2 in the cell transformants of a Sec2 tester strain were streaked to 5-FOA media to select for loss of the The data set supporting the results of this article are available in the Protein Data Bank (PDB) repository Accession Codes 4ZDW and 4Z8Y.wt and Sec4pV29 we performed nucleotide dissociation experiments using standard procedures [V29 was observed to be 6.5 × 10−5 s−1 . The rates obtained for the intrinsic nucleotide dissociation of Sec4p were very similar to those calculated for Rab3a [−5 s−1 which is 4 fold slower than the 2.2 × 10−4 s−1 calculated for the wild type protein [V29 at a significantly slower rate. In addition, the two curves present a noticeable difference in amplitude, which we interpret as being a result of partial dissociation of mantGDP nucleotide from Sec4pV29. Previous work has shown that Sec2p stably associates with the GDP bound form of Sec4pV29 but exclusively with the free form of Sec4pwt 30. We hypothesized that Sec2p is able to bind and change the conformation of both switch regions of Sec4pV29 in a similar fashion to that of the wild type protein (confirmed by the ternary complex Sec4pV29.GDP.Sec2p). However the bound nucleotide does not completely dissociate from its complex with Sec4pV29, which could be explained by the decrease in the nucleotide dissociation observed in our experiments  not considering the switch regions (RMSD 1.2 Å for 159 Cα atoms). Some of the crystal contacts are made through interactions with zinc atoms that are coordinated by Asp43 and Asp166 from molecule B, and His121 from molecule A. Besides the presence of the mutation S29V, the P-loop region from Sec4pV29.GDP and Sec4pwt.GDP [V29.GDP contains two molecules in the asymmetric unit (chains A and B). The switch I region is disordered in the molecule A and ordered in molecule B. The SWI of molecule B adopts a different conformation from that observed on the Sec4pwt.GDP structure. However, this new conformation is held in place by interactions with symmetric related molecules and is probably an artifact of crystal packing. The switch II region for molecule A presents the same conformation observed on molecule C of the Sec4pwt.GDP structure [wt.GDP was the interaction between the Arg81 of the SWII and the Ser29 of the P-loop. The side chain of the residue Arg81 forms a 2.8 Å length hydrogen bond with the side chain of the Ser29 residue. This interaction is not observed in the Sec4pV29.GDP structure . The wild type Sec4p can only stably associate with Sec2p in the absence of magnesium [V29 and Sec2p. This enabled us to obtain crystals for the ternary complex Sec4pV29.GDP.Sec2p and solve the structure at 2.9 Å resolution  which reflects the high flexibility of the complex [V29.GDP.Sec2p shares high similarity with the Sec4p.Sec2p complex bound to phosphate [V29.GDP.Sec2p and Sec4p.Phosphate.Sec2p structures is performed using only the Cα atoms of the Sec4p binding site of Sec2p (residues 100–120), it is clear that the mutant active site is in a more closed conformation than the wild type  and 153–157 (NLKKV). In the first construct alanines substituted a total of three leucine residues , two of which participate in the coiled-coil packing. In the second construct only the Leu154 participates in the coiled coil formation. As expected the mutant Sec2pAla142–147 shows a large reduction in the nucleotide dissociation with a rate similar to the truncation Sec2p51–142 . Besides conformational differences observed by the COOH-terminal of Sec2p, there are no major differences in the interaction interface between Sec4p and Sec2p between both structures. The second hypothesis is based on the fact that Sec2p forms a dimer and the GEF activity depends on the dimerization of the two monomeric helices. We could expect that disturbing the dimer formation of Sec2p would lead to loss of GEF activity. Since the region 142–160 presents an important dimerization interface, we hypothesized that the deletion of this region as seen in the truncation Sec251–142 would lead to a weaker interaction between the two monomers of Sec2p, directly reflecting a decrease in the dissociation constant for the dimer. To address this question we designed mutants in which a cysteine residue from one monomer would be able to interact with the symmetric residue from the other monomer and form a disulfide bond to generate a stable dimer. We designed two mutants, a single mutant on the COOH-terminal of Sec2p51–142 (K140C) and a double mutant where both upstream and downstream regions of the Sec4p binding site were mutated (A91C and K140C). The double mutant of Sec2p generated insoluble protein and was not used in further experiments. The single mutant Sec2S1 ‐ 142K140C rendered a very soluble and stable protein and was used for the nucleotide dissociation assays. The mutation K140C dramatically increased the GEF activity of the truncation Sec2p51–142 . These data suggest that the formation of a disulfide bond between the two monomers of Sec2p increases the affinity constant and the increase in protein activity reflects enhanced dimer stability. Treatment with a reducing agent (DTT) brings the activity of the mutant Sec2S1 ‐ 142K140C back close to its normal levels observed with Sec2p51–182  reveals that besides the overall similarities, a small shift in the P-loop results in a slightly tighter active site. The distance between the P-loop and Switch I, more specifically between the residues Val29 and Ile50 is shortened in comparison with the structure Sec4pwt.phosphate.Sec2p. The slight alteration in the active site, places the Ile50 residue very close to the magnesium binding site. It is thought that the Ile50 residue plays a critical role preventing magnesium rebinding to the protein after complex formation [V29 can form a stable complex with Sec2p.The difference in nucleotide dissociation rate is most likely explained by changes in the protein-nucleotide environment created by the hydrophobic side chain of the Val29. The mutation from the polar residue serine to the just slightly larger hydrophobic valine creates a more hydrophobic local environment. We can predict that the energy cost to solvate the active site of the mutant Val29 would be larger than for the wild type protein and could indirectly result in the increased binding affinity of the protein for the nucleotide. Furthermore, Sec2p stimulates the nucleotide dissociation from Sec4p complex  and the  complex , and tog51–142 stimulates nucleotide release from Sec4p at a much slower rate than the truncation containing the 18 residues in region 142–160. This was somehow surprising because the Sec4p binding site of Sec2p lies between residues 100–120, and the absence of the region 142–160 would not directly affect the protein protein interaction. However, looking closely, this region possesses several leucine residues forming a classical coiled-coil packing interface that could be important for dimer formation and protein stability. Mutation of the leucine residues participating in the coiled coil interaction to alanines reduced protein activity, most likely through a disruption of the dimer coiled-coil structure.During our nucleotide dissociation studies we discovered that the truncation Sec2p51–142. This truncation of Sec2p contains all the elements necessary for GEF activity however, has a slower activity than a protein which contains the dimer interface region between residues 142–160. The idea is that the formation of a disulfide linkage between the equivalent cysteine residues from both monomers would recapitulate dimerization in the absence of residues 142–160. The results of the nucleotide exchange assays show that in the absence of reducing agent the nucleotide dissociation rate for Sec2S1 ‐ 142K140C increased drastically in comparison to the wild type truncation Sec2p51–142, to a level similar to the truncation containing the coiled coil region 142–160 of Sec2p. These data indicates that although the region 142–160 is not directly involved in the binding to Sec4p this region is important for protein dimerization and indirectly affects protein activity. In addition, we determined that the first 50 residues in the Sec2p NH2-terminus do not affect GEF activity, but are important for full Sec2p function in vivo. This study extends the determination of a minimal GEF region for Sec4 and Sec2p interaction and establishes a minimal region of Sec2p containing the fully active enzyme, which we determined to be between residues 51–160. The intrinsic nucleotide dissociation rate of Sec4p is high relative to other Ras-related GTPases and the biological necessity for the potent GEF activity of Sec2p remains a mystery. A complete understanding of the mechanism of Sec2p action, together with functional analysis and interactions with other protein partners will provide important insights into the role of Sec4p and related Rab GTPases during exocytosis.To further validate the idea that disruption of dimerization causes reduced Sec2p activity, we created the mutant K140C on the truncation of Sec2p


An experiment (CLOUD7 campaign) was set up at the CLOUD (Cosmics Leaving OUtdoor Droplets) chamber to investigate the quantitative detection of H2SO4 in the presence of dimethylamine by CIMS at atmospherically relevant concentrations. For the first time in the CLOUD experiment, the monomer sulfuric acid concentration was measured by a CIMS and by two CI‐APi‐TOF  mass spectrometers. In addition, neutral sulfuric acid clusters were measured with the CI‐APi‐TOFs. The CLOUD7 measurements show that in the presence of dimethylamine (<5 to 70 pptv) the sulfuric acid monomer measured by the CIMS represents only a fraction of the total H2SO4, contained in the monomer and the clusters that is available for particle growth. Although it was found that the addition of dimethylamine dramatically changes the H2SO4 cluster distribution compared to binary (H2SO4‐H2O) conditions, the CIMS detection efficiency does not seem to depend substantially on whether an individual H2SO4 monomer is clustered with a DMA molecule. The experimental observations are supported by numerical simulations based on A Self‐contained Atmospheric chemistry coDe coupled with a molecular process model (Sulfuric Acid Water NUCleation) operated in the kinetic limit.Sulfuric acid is widely recognized as a very important substance driving atmospheric aerosol nucleation. Based on quantum chemical calculations it has been suggested that the quantitative detection of gas phase sulfuric acid (H 2SO4 concentration measured by CIMS in CLOUD was influenced by the presence of dimethylamineHCIMS's detection efficiency is not substantially influenced by the presence of dimethylamineInformation of sulfuric acid cluster composition by CI‐APi‐TOF explains most of the observed effect The climate forcing due to aerosol‐climate interaction causes the largest source of uncertainty in present climate models [2SO4) [Kulmala et al., Riipinen et al., Kirkby et al., 2SO4‐H2O system along with ammonia [Benson et al., Kirkby et al., Zhang et al., Metzger et al., Riccobono et al., Schobesberger et al., Kurtén et al., Almeida et al., Kürten et al., 2SO4‐H2O system, from the addition of atmospherically relevant concentrations of ammonia [Kirkby et al., Kirkby et al., One of the primary vapors responsible for nucleation is sulfuric acid  could be influenced by the presence of amines , (CH3)2NH). In this way, the clustering between H2SO4 and (CH3)2NH leading to a H2SO4·(CH3)2NH complex coupled to H2O molecules, abbreviated as SA·DMA in the following, could affect interpretation of the relationship between the derived sulfuric acid monomer concentration, the nucleation rate, and the growth rate [Kurtén et al., Kupiainen‐Määttä et al., Loukonen et al., Kurtén et al. [·DMA and the CIMS reagent ion (NO3−) could be considerably slower than the reaction rate between the bare H2SO4 monomer and the nitrate ion based on quantum chemistry computations of proton transfer energies. On the other hand, a more recent quantum chemistry study by Kupiainen‐Määttä et al. [·DMA cluster can be ionized by the nitrate ion at the collision limit. However, due to the higher dipole moment of SA·DMA compared to H2SO4, the collision rate with the reagent ion could be even somewhat higher for the sulfuric acid amine cluster. Clustering of the sulfuric acid with amines would therefore tend to increase their detection probability, instead of decreasing it, as it was suggested previously. As the SA·DMA clusters will participate in the nucleation even more efficiently than the bare H2SO4 molecules [Kürten et al., 2SO4 and SA·DMA).In addition to laboratory experiments, theoretical studies using quantum chemical calculations have also focused on the role of amines in aerosol nucleation [n et al. , the reaä et al.  which in2SO4 molecule or clusters containing 1 SA but potentially also water molecules or amine molecules, with the term amine in this study being used for dimethylamine. Similarly, the dimer contains 2 SA molecules, the trimer 3 SA, etc., but with varying amounts of water or amine. A recent study by Neitola et al. [2SO4‐H2O‐(CH3)2NH system investigated at CLOUD, it was found that the measured particle growth rates exceeded the expected growth rate due to condensation of sulfuric acid monomers by about a factor of 10 . This strongly enhanced growth could, however, not be explained merely by the presence of SA·DMA but also required a substantial amount of sulfuric acid contained in clusters (dimers and larger) contributing to the growth.Here we define SA monomers as either the single Ha et al.  showed aKürten et al. [Based on the findings by K. Lehtipalo et al.  and the earlier work by n et al.  this stu2This study presents measurements that were conducted in the CLOUD chamber at CERN between October and December 2012 (CLOUD7 campaign).2.13 chamber with unique features [Kirkby et al., Duplissy et al., 3, SO2, and amine) are introduced via independent gas lines. Typical condition of the chamber is at a pressure slightly higher than ambient atmospheric pressure, at a constant temperature of 278.15 K and at a relative humidity (RH) of 38%. The homogeneous distribution of chamber air is established by a pair of mixing fans [Voigtländer et al., Kupc et al., 2SO4 is then formed in situ from the reaction of SO2 with OH [Kupc et al., Nucleation experiments under atmospherically relevant conditions were conducted at the CLOUD (Cosmics Leaving OUtdoor Droplets) aerosol chamber. The CLOUD chamber is an electropolished stainless steel, 26.1 m+), creating variable levels of ion concentrations in the chamber [Duplissy et al., Kirkby et al., Almeida et al., Kürten et al., Praplan et al., A focus of the CLOUD measurements is to distinguish aerosol nucleation processes initiated with and without the aid of charged molecules. The CERN Proton Synchrotron can be used to expose the CLOUD chamber to a diverged beam of 3.5 GeV/c pions  is widely used both in laboratory and field experiments for the very sensitive detection of gaseous H2SO4 is thereby converted to bisulfate (HSO4−) through the reaction with nitrate primary ions (NO3−(HNO3)k = 0–2). These reagent ions are produced from a negative corona discharge when nitric acid is added to the sheath gas of the CI part of the instrument [Kürten et al., Rondo et al., ·DMA clusters produced in the CLOUD chamber are ionized via the following reactions:The working principle of the CIMS is to selectively charge sulfuric acid in the sample gas by means of chemical ionization and then detect the product ions with a mass spectrometer. A small fraction of the H·DMA cluster is ionized [Ortega et al., 4−, and most reagent ions are likewise converted to NO3−. While we expect that DMA already evaporates from the sulfuric acid monomer ions in the flow tube, due to the use of the CDC, any remaining DMA will be stripped off and any direct detection of DMA associated with a sulfuric acid monomer is not possible. Since the sulfuric acid monomer concentration is evaluated from the count rates of HSO4− and NO3−, it includes the contribution from H2SO4 as well as from SA·DMA. Regarding the charging efficiency of the sampled SA·DMA clusters, there is yet no experimental evidence but only theoretical studies supporting either lower [Kurtén et al., Kupiainen‐Määttä et al., 2SO4 molecules. However, if the second reaction 4− would change significantly upon the amine injection.All species of reaction 2.3Kürten et al., Jokinen et al., Kürten et al., 4− and NO3− [Kürten et al., 3 does not evaporate from the newly formed charged cluster, the DMA would be stabilized in the SA·DMA cluster as shown via the following reactions:A detailed insight into the elemental composition of molecular clusters in the sulfuric acid, water, and amine system is provided by a Chemical Ionization‐Atmospheric Pressure interface‐Time of Flight (CI‐APi‐TOF) mass spectrometer [1), dimer (N2), trimer (N3), tetramer (N4), and pentamer (N5) refer to the number of sulfuric acid molecules in the cluster coupled with H2O and amine molecules. To obtain the cluster concentrations (N2 to N5), the signals with different amounts of amine but with a certain number of sulfur atoms are added up [Kürten et al., 4 cm−3 can be reached for clusters containing four or five sulfuric acid and several amine molecules [Kürten et al., Kürten et al., In our study monomer sulfuric acid  model for atmospheric chemistry [C is determined so that the wall loss of H2SO4 agrees with the wall loss rate constant of 1.7 × 10−3 s−1 given in Almeida et al. [D of H2SO4 is taken from Hanson and Eisele [iD, of other small chemical species  are estimated from kinetic gas theory. The trace gases O3, SO2, and H2O are considered to be in equilibrium with the wall.a et al. , while td Eisele . Diffusiij, is implemented as function of the UV light aperture and is proportional to the UV light power as given in Kupc et al. [The UV light photolysis rate, c et al. (2)ji=a×jth, for production of O(1D) and O(3P) from O3 as well as the photolysis of H2O2 to OH was calculated based on the UV light spectrum while factor a is a parameter to adjust the photolysis rate to the UV light in the chamber. The necessity of this factor relies on the fact that the actual photon flux in the chamber is not known and has to be determined for each CLOUD campaign due to changes in the UV lamps, such as aging or replacement. For this study the photolysis rate was adjusted based on fits to experiments without any amine present. The polynomial relationship between UV aperture (per cent) and UV intensity (per cent) represented as Pap is defined in Kupc et al. [1D) and O(3P) and OH are 2.3 × 10−6, 2.7 × 10−7, and 2.5 × 10−8, respectively.The theoretical UV light photolysis rate, c et al.  and tran2.4.2Lovejoy et al., ·DMA cluster [Ehrhart and Curtius, Kürten et al., Ehrhart and Curtius, 2SO4 and one amine was used for the simulations; the ratio of 1:1 was also assumed for the clusters and particles. The density of H2SO4‐DMA was set to 1400 kg m−3 [Qiu and Zhang, 2, O3, and H2O, the temperature of the chamber, and the UV aperture of CLOUD's UV system. In Figure The SAWNUC (Sulfuric Acid Water NUCleation) model was originally developed for binary sulfuric acid‐water nucleation [2SO4, but diffusion coefficients were calculated from Cunningham‐corrected Stokes‐Einstein diffusion coefficients [Baron and Willeke, Wall loss of clusters was implemented in the same way as for H33.12SO4‐H2O system with other condensable compounds present only as trace impurities [Bianchi et al., 2SO4 concentrations from 1 × 106 to 1 × 108 molecule cm−3 was studied. The production of gaseous sulfuric acid is established by means of photolytic reactions of the trace gases O3 and SO2 initiated by the UV light illumination. Variation of the UV light intensity results in a proportional response in the sulfuric acid production rate, with its equilibrium concentration being established within 650 ± 33 s when the production rate is balanced by the wall loss rate. An overview of a typical binary experimental run is given in Figure 2SO4 concentration for different UV light aperture settings. The measured sulfuric acid monomer concentrations show an almost linear dependence on the applied UV light intensity for the binary system  was added to the CLOUD chamber after equilibrium of the sulfuric acid concentration was reached for binary conditions  .Apart from the decrease of the measured monomer sulfuric acid concentrations, we also observed a change of the characteristic time period of the Hy Figure . The diskR1 ≠ kR2) [Kurtén et al., Kupiainen‐Määttä et al., Hanson and Eisele, ·DMA cluster can bind to water molecules and its effective evaporation rate decreases [Olenius et al., 2SO4 from reaction with SO2, are partly consumed due to the added amine. The reaction rate constant between OH and dimethylamine (6.5 × 10−11 cm3 molecule−1 s−1 at 298 K) is approximately 7 times larger than the one between OH and SO2 {9 × 10−12 cm3 molecule−1 s−1 at 298 K [Atkinson et al., 2 during CLOUD7 was 60 ppbv compared to a maximum of ~70 pptv of amine. Therefore, the expected change in the produced H2SO4 due to the amine addition should be less than 0.5%. The possibility of a significant difference of the ion molecule reaction rate constants kR1 and kR2 will be discussed below.Regarding the alteration of the sulfuric acid‐water system due to the introduction of amine, we should take into consideration the potential contribution of several other effects in addition to the formation of the larger clusters: change of diffusivity of the monomers, OH consumption, or any change of the ion‐molecule‐reaction rate in the CIMS‐reactor (3.22SO4 monomer is being transferred into larger SAxDMAy (x > 1) clusters that do not evaporate rapidly, confirming the stabilizing role of the amine. A recent study by Kürten et al. [2SO4 and 16 DMA molecules confirming the important role of amines for the formation of stable clusters by nucleation with sulfuric acid. In addition, as displayed in Figure While the measurements show a decrease of the sulfuric acid monomer concentration, the CI‐APi‐TOF's cluster measurements reveal information on cluster formation that explains the observed reduction of the monomer to a large degree. As shown in Figure n et al.  reportedtotal sulfuric acid, SAtotal by summing up the SA molecules contained in the monomers and clusters up to the pentamer; totalSA = ∑i=15iiN  along wKürten et al., Although there are even larger clusters, we define the total sulfuric acid in this study as the sulfuric acid which is contained in clusters up to the pentamer because this was the largest size for which the concentrations were quantitatively derived [i‐mer with another SA i‐mer leads to a further clustering while it is assumed that no evaporation of clusters occurs. Indeed, the comparison of the modeled (red curve) and measured total sulfuric acid cluster concentration (red dots) reveals that there is still a discrepancy between the measured total sulfuric acid and the amount expected for the kinetic limit. The discrepancy between the modeled and measured total sulfuric acid indicates either a declining transmission of the mass spectrometer for larger clusters or a decrease of the ion‐molecule reaction rate constants with increasing cluster size. Most likely, these effects can explain the remaining discrepancy between the kinetic limit calculation and the observed total sulfuric acid. Regarding the magnitude of the discrepancy between total sulfuric acid and sulfuric acid monomer, it is also important to note that it can be even larger than what is shown in Figure ·DMA cluster in the atmosphere is a function of the conditions and will depend on the cluster distribution under specific conditions, taking into consideration, for example, losses to preexisting particles. A more detailed study regarding the contribution of clustering to the nanoparticle growth over a wide range of atmospheric conditions is included in the submitted study of K. Lehtipalo et al. .In order to investigate quantitatively the magnitude of the “hidden” sulfuric acid that contributes to coagulation and growth but is not measured by the CIMS, our experimental measurements were compared with theoretical predictions Figure . All pos43) and dimethylamine (DMA) leading to the formation of larger clusters and particles has been demonstrated in earlier studies [Kirkby et al., Almeida et al., Bianchi et al., Kürten et al., 2SO4‐H2O‐NH3 system, the nature of the chemical bond of sulfuric acid to dimethylamine reveals that the H2SO4‐H2O‐DMA system forms structures that are significantly more strongly bound [Loukonen et al., Kürten et al., 2SO4 monomer concentrations in the atmosphere and in the laboratory. It has been suspected that the efficiency of the ion‐molecule‐reaction that is used for the detection of the H2SO4 monomers changes in the presence of dimethylamine [Kurtén et al., Kupiainen‐Määttä et al., 2SO4 monomers in the absence and presence of dimethylamine to see if the monomer signals change either (a) due to the onset of highly efficient particle nucleation and formation of clusters which reduce the monomer signal or (b) because the rate constant kR2 significantly differs from kR1.The capability of sulfuric acid molecules to bind with base molecules such as ammonia . Distinct differences were observed, from both CIMS and CI‐APi‐TOF, between binary H2SO4‐H2O and ternary H2SO4‐H2O‐DMA nucleation experiments. It was found that for the H2SO4‐H2O‐DMA ternary system, the sulfuric acid monomer concentration as measured by the bisulfate ion (m/z 97) was influenced by the presence of dimethylamine. We measured monomer sulfuric acid concentration using both CIMS and CI‐APi‐TOF mass spectrometers and observed a reduction of the monomer signal after DMA was introduced. In addition, a simultaneous increase of all the larger sulfuric acid clusters (dimer to pentamer) was observed. Therefore, the linear dependency of the monomer sulfuric acid to the UV light intensity that applies to the binary system does not hold for the amine ternary system anymore. Instead, a close to square root dependency is observed. From the binary and DMA ternary experiments it was found that the time required for the H2SO4 monomer concentration to reach its equilibrium is lowered in the presence of amine by a factor of ~4, although the production rate of sulfuric acid remained constant. This is explained by the increased monomer sink due to efficient formation of dimers and collisions with larger clusters. It is suggested that effects such as changing diffusivity of the clusters, OH consumption due to reaction with amine, and changing ion molecule reaction rate after the introduction of amine cause only a minor contribution to the change of the monomer signal. The CI‐APi‐TOF measurements do provide information on cluster composition that explains most of the observed effect.Hi‐mer that collides with another sulfuric acid i‐mer sticks to that cluster. A remaining difference is most likely contributed to reduced transmission of the larger clusters in the CI‐APi‐TOF and potentially a smaller ion‐molecule reaction rate constant for the higher i‐mers compared to the monomer. In addition, the obtained results of sulfuric acid monomer and its clusters from the chemical ionization mass analyzers are in good agreement with the performed SAWNUC‐ASAD chemical model.The experimental results are in good agreement with modeling studies when assuming that the base‐stabilization by amine leads to kinetically controlled growth of the sulfuric acid clusters where every sulfuric acid 2SO4 are correctly measured by the CIMS technique regardless of the presence of amine, therefore not supporting suggested changes of the charging efficiency compared with the bare H2SO4 molecules as proposed by Kurtén et al. [Kupiainen‐Määttä et al. [2SO4 to the growth, ideally the dimers and larger clusters have to be measured quantitatively as well to determine the amount of sulfuric acid “hidden” in these clusters and the total sulfuric acid. Furthermore, substances like dimethylamine which are able to shift a significant fraction of the monomers into larger i‐mers have to be determined along with the H2SO4 measurements.A major conclusion of our study is that the monomer concentrations of Hn et al.  or Kupiaä et al. . But theFigure S1 and Table S1Click here for additional data file.