IR spectroelectrochemical study of the binding of carbon monoxide to the active site of Desulfovibrio fructosovorans Ni-Fe hydrogenase
25 páginas, 9 figuras y 2 tablas ; The binding of carbon monoxide a competitive inhibitor of many hydrogenases, to the active site of Desulfovibrio fructosovorans hydrogenase has been studied by infrared spectroscopy in a spectroelectrochemical cell. Direct evidence has been obtained of which redox states of the enzyme can bind extrinsic CO. Redox states A, B and SU do not bind extrinsic CO. only after reductive activation of the hydrogenase can CO bind to the active site. Two states with bound extrinsic CO can be distinguished by FTIR. These two states are in redox equilibrium and are most probably due to different oxidation states of the proximal 4Fe-4S cluster. Vibrational frequencies and theoretical quantum mechanics studies (DFT) of this process preclude the possibility of strong bonding of extrinsic CO to the Fe or Ni atoms or the site. We propose that CO inhibition is caused by weak interaction of the extrinsic ligand with the Ni atom, blocking electron and proton transfer at the active site. A calculated structure with a weakly bound extrinsic CO at Ni has relative CO frequencies in excellent agreement with the experimental ones. ; The Spanish Ministry of Science and Technology (project BQU2000-0991) and the European Union BIOTECH program (grant BIO-98-0280) supported this work. We thank Prof. Siem Albracht for useful discussions, Dr. Arturo Martinez-Arias for doing the EPR measurements, the Autonomous Community of Madrid (CAM) for the postdoctoral fellowship of A. L. De Lacey and the German Academic Exchange Service (DAAD) for the postdoctoral fellowship of C. Stadler. MBH thanks NSF (CHE-9800184) for financial support. ; Peer reviewed
