This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.The bacteria R. rubrum (Rr) and C. hydrogenoformans (Ch) contain a Ni carbon monoxide dehydrogenase (CODH) that enables them to grow on CO as their sole carbon source. The active site of Ni CODH is the so-called 'C-cluster' a novel Ni-Fe-S center that catalyzes the oxidation of CO to CO2. Recent X-ray crystallography has shown the 'C-cluster' to comprise a Ni coordinated to a FeS4 unit, bridged to an external Fe site. This was a surprise, as prior EXAFS analysis had excluded a symmetric NiFeS4 unit. However, the various 'C-cluster' crystal structures show significant differences with each other, and, they also show evidence of cluster heterogeneity within a given crystal. We have recent EXAFS data that shows evidence of cluster re-arrangement on reduction and/or CO binding. The implication is that the cluster structure dynamically rearranges, either as part of an 'activation' process, or as part of its catalytic mechanism. We propose to use EXAFS to investigate the structural changes of the 'C-cluster' sites for Rr and Ch CODH. This is part of a comprehensive spectroscopic study on the C-cluster. We plan to study enzyme that has been: reductively poised; CO-bound and inhibited. We have available variants of Rr CODH where active site residues have been modified, one where the external Fe is deleted. We also have Co-CODH and Zn-CODH, where the Ni has been replaced by another transition metal. We are confident that, using other spectroscopies such as EPR, FT-IR and resonance Raman as controls, we can prepare homogeneous C-cluster samples for EXAFS analysis. This should enable us to parameterize the C-cluster structural dynamics and from there understand the underpinning cluster chemistry and eventually the catalytic cycle of this biologically important enzyme system. In addition, the novel bioinorganic chemistry uncovered could well lead to the development of novel catalysts.

National Institute of Health (NIH)
National Center for Research Resources (NCRR)
Biotechnology Resource Grants (P41)
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