Antibiotic resistance has emerged as a serious threat to public health. Much of the resistance is found in the widely used beta-lactam antibiotic groups which include penicillins and carbapenems. This resistance is thought to be due to genetic mutations that cause the development of an enzyme called metallo-beta-lactamase (MBL). MBLs include two zinc ions at the active site which prove to be problematic in inhibition. Unavailability of therapeutic inhibitors has led to a search for novel inhibitors. The MBL from Bacillus anthracis (Bla2) was chosen for this study. Successful purification of Bla2 using a Ni2+ column was confirmed by SDS-PAGE analysis, and attempts were made to inhibit this enzyme using hydroxamate-group-containing molecules. After obtaining sufficient amounts of Bla2, attempts were made at inhibiting the enzyme using hydroxamate-group-containing molecules. Initial kinetic studies suggest significant inhibition in a molecule containing two hydroxamate functional groups with a promising IC50 value but no significant inhibition in the single hydroxamate containing molecule. Later kinetic analysis reveals the significant inhibition to be competitive with an extraordinary Ki value. Dimethyl sulfoxide was used to increase inhibitor solubility, and further enzyme assay revealed enzyme stability up to approximately 2% dimethyl sulfoxide. In silico analysis showed inhibitor docking to the active site of Bla2 which provides a link between experimental and theoretical work. These findings offer new avenues for designing better inhibitors against MBLs.

Inhibition testing of the metallo-beta-lactamase (Bla2) found in antibiotic resistant Bacillus anthracis

252nd National Meeting (2016)

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