title: Recognition of a cysteine substrate by E. coli gamma-glutamylcysteine synthetase probed by sulfoximine-based transition-state analogue inhibitors.
abstract: A series of sulfoximine-based transition-state analogue inhibitors with a varying alkyl side chain was synthesized to probe the recognition of a Cys substrate by E. coli gamma-glutamylcysteine synthetase (gamma-GCS). The sulfoximines with a small alkyl group (H, methyl, ethyl, propyl, butyl and CH2OH) each served as a slow-binding inhibitor, the sulfoximine with an ethyl being by far the most potent inhibitor to cause facile and irreversible enzyme inhibition. As the size of the side chain changed from an ethyl, the inhibition potency markedly decreased to reduce the overall affinity with concomitant loss in the inactivation rate and with facile enzyme reactivation by dilution. The sulfoximine without a side chain inhibited the enzyme with almost the same potency as that of L-buthionine-(SR)-sulfoximine (BSO). The free energy difference calculated from the inhibition constants indicates that the side chain of Cys was recognized by its size through hydrophobic interaction and contributed almost equally or even more than the carboxy group to the overall binding of Cys in the transition state.
authors.affiliation: Institute for Chemical Research, Kyoto University, Uji, Japan. hiratake@scl.kyoto-u.ac.jp
authors.forename: Jun
authors.initials: J
authors.lastname: Hiratake
authors.forename: Takayuki
authors.initials: T
authors.lastname: Irie
authors.forename: Nobuya
authors.initials: N
authors.lastname: Tokutake
authors.forename: Jun'ichi
authors.initials: J
authors.lastname: Oda
mesh_terms: D019328:Buthionine Sulfoximine / Q000494:pharmacology*; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000494:pharmacology*; D004926:Escherichia coli / Q000201:enzymology*; D005721:Glutamate-Cysteine Ligase / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D007202:Indicators and Reagents; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity
chemical_list: D004791:Enzyme Inhibitors; D007202:Indicators and Reagents; D019328:Buthionine Sulfoximine; D005721:Glutamate-Cysteine Ligase; D003545:Cysteine