engleski

Conformational changes accompanying aminoacyl adenylate formation by peptide synthetases

Peptide synthetases constitute a family of multifunctional enzyme systems involved in the non-ribosomal biosynthesis of a variety of microbial peptide products, some with important medical and biotechnological applications as antibiotics (gramicidin S, tyrocidine, penicillins), immunomodulators (cyclosporin), extracellular surfactans (surfactin), and sideophores (enterobactin). The modular construction of the polypeptide chain opens the possibility for manipulation, and consequently generation of novel peptides with modified biological activity. Substrate activation is accomplished by formation of an acyladenylate, followed by charging of the activated moiety onto the sulfhydryl group of the 4˘- phosphopantetheine cofactor for subsequent modification and elongation. The crystal structure has provided a wealth of structural information, however the substrate binding sites, and the conformational changes which occur upon binding and prior to product release are currently unresolved issues. The major unanswered questions about the dynamic behaviour of peptide synthetases regard the binding of ATP, the juxtaposition of the b- and g-phosphates for adenylate formation, the mechanism of PPi release, and the significance of the thioesterification site in regulating the conformational stability of the activated intermediates. The functional importance of the two flexible loops comprising the most conserved residues, Arg416 and Lys186, has been studied by the use of mutant analysis, limited proteolysis, and chemical modification of tyrocidine synthetase 1. It has been proposed that the (S, T)GT(T, S)GxPKG signature sequence is involved in orienting the b- and g- phosphates for the catalytic reaction. The 4˘- phosphopantetheine cofactor is likely to be relevant for maintainance of the active site conformation, and consequently protection from decomposition by hydrolysis.

Conformational changes; Peptide synthetase

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