Format of Original
American Chemical Society
Journal of the American Chemical Society
Original Item ID
In an effort to probe the structure, mechanism, and biochemical properties of metallo-β-lactamase Bla2 from Bacillus anthracis, the enzyme was overexpressed, purified, and characterized. Metal analyses demonstrated that recombinant Bla2 tightly binds 1 equiv of Zn(II). Steady-state kinetic studies showed that mono-Zn(II) Bla2 (1Zn-Bla2) is active, while di-Zn(II) Bla2 (ZnZn-Bla2) was unstable. Catalytically, 1Zn-Bla2 behaves like the related enzymes CcrA and L1. In contrast, di-Co(II) Bla2 (CoCo-Bla2) is substantially more active than the mono-Co(II) analogue. Rapid kinetics and UV−vis, 1H NMR, EPR, and EXAFS spectroscopic studies show that Co(II) binding to Bla2 is distributed, while EXAFS shows that Zn(II) binding is sequential. To our knowledge, this is the first documented example of a Zn enzyme that binds Co(II) and Zn(II) via distinct mechanisms, underscoring the need to demonstrate transferability when extrapolating results on Co(II)-substituted proteins to the native Zn(II)-containing forms.
Hawk, Megan J.; Breece, Robert M.; Hajdin, Christine E.; Bender, Katherine M.; Hu, Zhenxin; Costello, Alison L.; Bennett, Brian; Tierney, David L.; and Crowder, Michael W., "Differential Binding of Co(II) and Zn(II) to Metallo-β-Lactamase Bla2 from Bacillus anthracis" (2009). Physics Faculty Research and Publications. 16.