Physisorption of enzymatically active chymotrypsin on titania colloidal particles.
In this study we use a straightforward experimental method to probe the presence and activity of the proteolytic enzyme α-chymotrypsin adsorbed on titania colloidal particles. We show that the adsorption of α-chymotrypsin on the particles is irreversible and pH-dependent. At pH 8 the amount of adsorbed chymotrypsin is threefold higher compared to the adsorption at pH 5. However, we observe that the adsorption is accompanied by a substantial loss of enzymatic activity, and only around 6-9% of the initial enzyme activity is retained. A Michaelis-Menten kinetics analysis of both unbound and TiO2-bound chymotrypsin shows that the K(M) value is increased from ∼10 μM for free chymotrypsin to ∼40 μM for the particle bound enzyme. Such activity decrease could be related by the hindered accessibility of substrate to the active site of adsorbed chymotrypsin, or by adsorption-induced structural changes. Our simple experimental method does not require any complex technical equipment, can be applied to a broad range of hydrolytic enzymes and to various types of colloidal materials. Our approach allows an easy, fast and reliable determination of particle surface-bound enzyme activity and has high potential for development of future enzyme-based biotechnological and industrial processes.
['Derr|Ludmilla|L|', 'Dringen|Ralf|R|', 'Treccani|Laura|L|', 'Hildebrand|Nils|N|', 'Ciacchi|Lucio Colombi|LC|', 'Rezwan|Kurosch|K|']
D000327:Adsorption D020134:Catalytic Domain D002918:Chymotrypsin D003102:Colloids D057075:Enzyme Assays D004800:Enzymes, Immobilized D006863:Hydrogen-Ion Concentration D057927:Hydrophobic and Hydrophilic Interactions D007700:Kinetics D009596:Nitrophenols D014025:Titanium
Physisorption Enzymatic activity Chymotrypsin Colloidal titania particles Enzyme adsorption TiO(2)