1xql
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Effect of a Y265F Mutant on the Transamination Based Cycloserine Inactivation of Alanine Racemase
OverviewOverview
The requirement for d-alanine in the peptidoglycan layer of bacterial cell, walls is fulfilled in part by alanine racemase (EC 5.1.1.1), a pyridoxal, 5'-phosphate (PLP)-assisted enzyme. The enzyme utilizes two antiparallel, bases focused at the C(alpha) position and oriented perpendicular to the, PLP ring to facilitate the equilibration of alanine enantiomers., Understanding how this two-base system is utilized and controlled to yield, reaction specificity is therefore a potential means for designing, antibiotics. Cycloserine is a known alanine racemase suicide substrate, although its mechanism of inactivation is based on transaminase chemistry., Here we characterize the effects of a Y265F mutant (Tyr265 acts as the, catalytic base in the l-isomer case) of Bacillus stearothermophilus, alanine racemase on cycloserine inactivation. The Y265F mutant reduces, racemization activity 1600-fold [Watanabe, A., Yoshimura, T., Mikami, B., and Esaki, N. (1999) J. Biochem. 126, 781-786] and only leads to formation, of the isoxazole end product (the result of the transaminase pathway) in, the case of d-cycloserine, in contrast to results obtained using the, wild-type enzyme. l-Cycloserine, on the other hand, utilizes a number of, alternative pathways in the absence of Y265, emphasizing the importance of, Y265 in both the inactivation and racemization pathway. In combination, with the kinetics of inactivation, these results suggest roles for each of, the two catalytic bases in racemization and inactivation, as well as the, importance of Y265 in "steering" the chemistry to favor one pathway over, another.
About this StructureAbout this Structure
1XQL is a Single protein structure of sequence from Geobacillus stearothermophilus with PMP, PMH, PLP, 4AX and ACY as ligands. Active as Alanine racemase, with EC number 5.1.1.1 Full crystallographic information is available from OCA.
ReferenceReference
Effect of a Y265F mutant on the transamination-based cycloserine inactivation of alanine racemase., Fenn TD, Holyoak T, Stamper GF, Ringe D, Biochemistry. 2005 Apr 12;44(14):5317-27. PMID:15807525
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