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American Society for Biochemistry and Molecular Biology
Journal of Biological Chemistry
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In an effort to structurally probe the metal binding site in VanX, electronic absorption, EPR, and extended x-ray absorption fine structure (EXAFS) spectroscopic studies were conducted on Co(II)-substituted VanX. Electronic spectroscopy revealed the presence of Co(II) ligand field transitions that had molar absorptivities of ∼100 m–1 cm–1, which suggests that Co(II) is five-coordinate in Co(II)-substituted VanX. Low temperature EPR spectra of Co(II)-substituted VanX were simulated using spin Hamiltonian parameters of M = |±½〉, E/D = 0.14, greal(x,y) = 2.37, and grealS(z) = 2.03. These parameters lead to the prediction that Co(II) in the enzyme is five-coordinate and that there may be at least one solvent-derived ligand. Single scattering fits of EXAFS data indicate that the metal ions in both native Zn(II)-containing and Co(II)-substituted VanX have the same coordination number and that the metal ions are coordinated by 5 nitrogen/oxygen ligands at ∼ 2.0 Å. These data demonstrate that Co(II) (and Zn(II) from EXAFS studies) is five-coordinate in VanX in contrast to previous crystallographic studies (Bussiere, D. E., Pratt, S. D., Katz, L., Severin, J. M., Holzman, T., and Park, C. H. (1998) Mol. Cell 2, 75–84). These spectroscopic studies also demonstrate that the metal ion in Co(II)-substituted VanX when complexed with a phosphinate analog of substrate d-Ala-d-Ala is also five-coordinate.
Breece, Robert M.; Costello, Alison L.; Bennett, Brian; Sigdel, Tara K.; Matthews, Megan L.; Tierney, David L.; and Crowder, Michael W., "A Five-coordinate Metal Center in Co(II)-substituted VanX" (2005). Physics Faculty Research and Publications. 80.
ADA Accessible Version
Published version. Journal of Biological Chemistry, Vol. 280, No. 12 (March 25, 2005): 11074-11081. DOI. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc. Used with permission.
Brian Bennett was affiliated with the Medical College of Wisconsin at the time of publication.