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Inactivation of Glucosamine‐6‐Phosphate Synthase by N 3 ‐Oxoacyl Derivatives of L ‐2,3‐Diaminopropanoic Acid
Author(s) -
Jędrzejczak Robert,
Wojciechowski Marek,
Andruszkiewicz Ryszard,
Sowiński Paweł,
KotWasik Agata,
Milewski Sławomir
Publication year - 2012
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201100587
Subject(s) - chemistry , ketone , stereochemistry , enzyme , moiety , epoxide , active site , atp synthase , ligand (biochemistry) , double bond , affinity label , biochemistry , organic chemistry , catalysis , receptor
Abstract N 3 ‐Oxoacyl derivatives of L ‐2,3‐diaminopropanoic acid 1 – 4 , containing either an epoxide group or a conjugated double bond system, inactivate Saccharomyces cerevisiae glucosamine‐6‐phosphate (GlcN‐6‐P) synthase in a time‐ and concentration dependent manner. The results of kinetics studies on inactivation suggested a biphasic course, with formation of the enzyme–ligand complex preceding irreversible modification of the enzyme. The examined compounds differed markedly in their affinity to the enzyme active site. Inhibitors containing a phenyl ketone moiety bound much more strongly than their methyl ketone counterparts. The molecular mechanism of enzyme inactivation by phenyl ketone compounds 1 and 3 was elucidated by using a stepwise approach with 2D NMR, MS and UV–visible spectroscopy. A substituted thiazine derivative was identified as the final product of a model reaction between an epoxide compound, 1 , and L ‐cysteine ethyl ester (CEE); and the respective cyclic product, found as a result of reaction between 1 and CGIF tetrapeptide, was identical to the N‐terminal fragment of GlcN‐6‐P synthase. On the other hand, the reaction of a double‐bond‐containing compound, 3 , with CEE, CGIF and GlcN‐6‐P synthase led to the formation of a CS bond, without any further conversion or rearrangement. Molecular mechanisms of the reactions studied are proposed.