Effect of mutation of lysine-120, located at the entry to the active site of O-acetylserine sulfhydrylase-A from Salmonella typhimurium.
Biochim Biophys Acta
; 1784(4): 629-37, 2008 Apr.
Article
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| MEDLINE
| ID: mdl-18243146
ABSTRACT
O-Acetylserine sulfhydrylase catalyzes the final step of the biosynthesis of L-cysteine, the replacement of the beta-acetoxy group of O-acetyl-L-serine (OAS) by a thiol. The enzyme undergoes a conformational change to close the site upon formation of the external Schiff base (ESB) with OAS. Mutation of K120 to Q was predicted to destabilize the closed form of the ESB and decrease the rate. The K120Q mutant enzyme was prepared and characterized by UV-visible absorbance, fluorescence, visible CD, and 31P NMR spectral studies, as well as steady state and pre-steady state kinetic studies. Spectra suggest a shift in the tautomeric equilibrium toward the neutral enolimine and an increase in the rate of interconversion of the open and closed forms of the enzyme. A decrease in the rate of both half reactions likely reflects the stabilization of the ESB as a result of the increased rate of equilibration of the open and closed forms of the enzyme along the reaction pathway. Data suggest a role of K120 in helping to stabilize the closed conformation by participating in a new hydrogen bond to the backbone carbonyl of A231.
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Banco de datos:
MEDLINE
Asunto principal:
Salmonella typhimurium
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Proteínas Bacterianas
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Cisteína Sintasa
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Lisina
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Mutación
Idioma:
En
Revista:
Biochim Biophys Acta
Año:
2008
Tipo del documento:
Article