Analysis of the linker region joining the adenylation and carrier protein domains of the modular nonribosomal peptide synthetases.
Nonribosomal peptide synthetases (NRPSs) are multimodular proteins capable of producing important peptide natural products. Using an assembly line process, the amino acid substrate and peptide intermediates are passed between the active sites of different catalytic domains of the NRPS while bound covalently to a peptidyl carrier protein (PCP) domain. Examination of the linker sequences that join the NRPS adenylation and PCP domains identified several conserved proline residues that are not found in standalone adenylation domains. We examined the roles of these proline residues and neighboring conserved sequences through mutagenesis and biochemical analysis of the reaction catalyzed by the adenylation domain and the fully reconstituted NRPS pathway. In particular, we identified a conserved LPxP motif at the start of the adenylation-PCP linker. The LPxP motif interacts with a region on the adenylation domain to stabilize a critical catalytic lysine residue belonging to the A10 motif that immediately precedes the linker. Further, this interaction with the C-terminal subdomain of the adenylation domain may coordinate movement of the PCP with the conformational change of the adenylation domain. Through this work, we extend the conserved A10 motif of the adenylation domain and identify residues that enable proper adenylation domain function.
['Miller|Bradley R|BR|', 'Sundlov|Jesse A|JA|', 'Drake|Eric J|EJ|', 'Makin|Thomas A|TA|', 'Gulick|Andrew M|AM|']
D020816:Amino Acid Motifs D000595:Amino Acid Sequence D055162:Biocatalysis D002352:Carrier Proteins D019295:Computational Biology D017124:Conserved Sequence D030562:Databases, Protein D004758:Enterobactin D004926:Escherichia coli D029968:Escherichia coli Proteins D008239:Lysine D008958:Models, Molecular D009097:Multienzyme Complexes D050505:Mutant Proteins D010453:Peptide Synthases D011392:Proline D054730:Protein Interaction Domains and Motifs D011499:Protein Processing, Post-Translational D011994:Recombinant Proteins D012984:Software
acyl-carrier proteins enterobactin enzyme structure natural product biosynthesis secondary metabolism siderophores structural biology