2bfk
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BACILLUS CEREUS METALLO-BETA-LACTAMASE (BCII) ARG (121) CYS MUTANT. SOLVED AT PH7 USING 20MM ZNSO4 IN BUFFER. 1MM DTT WAS USED AS A REDUCING AGENT
OverviewOverview
The zinc-dependent metallo-beta-lactamases are a group of bacterial, enzymes that pose a threat to the future efficacy of present-day, antibiotics. Their mechanism is poorly understood, and there are no, clinically useful inhibitors. While most members of the group contain two, tightly bound zinc ions in their active sites, the Bacillus cereus enzyme, has a much lower affinity for its second zinc (Zn2), thought to be due to, the presence of Arg121 immediately beneath the floor of the active site, (cf. Cys/Ser/His121 in the bizinc enzymes). Crystal structures of the, Arg121Cys mutant of the B. cereus 569/H/9 enzyme were solved at pH 7.0, 5.0, and 4.5, each in the presence of either 20 microM or 20 mM Zn(2+) to, generate the mono- and bizinc forms, respectively. Surprisingly, the, structure of the active site was unaffected by the mutation; a network of, ordered water molecules replaced the interactions made by the arginine, side chain, and the occupancy of Zn2 appeared minimally changed. As the pH, was lowered, Zn2 moved away from one of its ligands, Asp120, but was, "tracked" by two others, Cys221 and His263. Furthermore, the hydroxide ion, (and proposed nucleophile for beta-lactam hydrolysis) was bound to Zn1 at, pH 5 and above but absent at pH 4.5. This provides experimental evidence, for an earlier proposed mechanism in which protonation of Asp120 and the, Zn1-bound hydroxide are the two events that lead to the loss of activity, at low pH.
About this StructureAbout this Structure
2BFK is a Single protein structure of sequence from Bacillus cereus with AZI, ZN, SO4 and GOL as ligands. Active as Beta-lactamase, with EC number 3.5.2.6 Structure known Active Site: AC1. Full crystallographic information is available from OCA.
ReferenceReference
Effect of pH on the active site of an Arg121Cys mutant of the metallo-beta-lactamase from Bacillus cereus: implications for the enzyme mechanism., Davies AM, Rasia RM, Vila AJ, Sutton BJ, Fabiane SM, Biochemistry. 2005 Mar 29;44(12):4841-9. PMID:15779910
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