CO Binding to the FeMo Cofactor of CO-Inhibited Nitrogenase: 13CO and 1H Q-Band ENDOR Investigation

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• 作者：Hong-In Lee ; Linda M. Cameron ; Brian J. Hales ; and Brian M. Hoffman
• 刊名：Journal of the American Chemical Society
• 出版年：1997
• 出版时间：October 22, 1997
• 年：1997
• 卷：119
• 期：42
• 页码：10121 - 10126
• 全文大小：154K
• 年卷期：v.119,no.42(October 22, 1997)
• ISSN：1520-5126

文摘

The resting state of nitrogenase shows an S= ³/₂ electron paramagnetic resonance (EPR)signal resultingfrom the FeMo-cofactor (MoFe₇S₉:homocitrate) ofthe MoFe protein. When the enzyme undergoes turnoverundera CO atmosphere, this signal disappears and two new ones appear: oneunder low pressure of CO (denoted lo-CO;0.08 atm) and the other under high pressure of CO (denoted hi-CO; 0.5atm). Our recent Q-band (35 GHz) ¹³C and⁵⁷Fe electron nuclear double resonance (ENDOR) studiesdemonstrated that one CO is bound to the FeMo-cofactorof lo-CO and two to the cofactor of hi-CO. [Christie, P. D.; Lee,H. I.; Cameron, L. M.; Hales, B. J.; Orme-Johnson, W. H.; Hoffman, B. M. J. Am.Chem. Soc. 1996, 118,8707-8709. Pollack, R. C.; Lee, H. I.; Cameron,L. M.; DeRose, V. J.; Hales, B. J.; Orme-Johnson, W. H.; Hoffman, B. M.J. Am. Chem. Soc.1995, 117, 8686-8687.] In the present report, we examine the CO-boundFeMo-cofactor in both the lo- and hi-CO forms of theMoFe protein from Azotobacter vinelandii by completeorientation-selective ¹³C and ¹H ENDORmeasurements.¹H ENDOR studies reveal that well-resolved signalsfrom a solvent-exchangeable proton seen in the restingstateFeMo-cofactor are lost in both of the CO-inhibited forms, indicating aloss in hydrogen bonding as compared to theresting state. This supports the hypothesis that CO binds near the"waist" of the cofactor. Determination of¹³Chyperfine tensors of bound ¹³CO to lo-CO and hi-COleads to the suggestion that the single CO bound to theFeMo-cofactor of lo-CO may bridge or semibridge two iron ions, while each ofthe two CO bound to hi-CO is a terminalligand. These ENDOR measurements and recent FTIR results ofThorneley and co-workers [George, S. J.; Ashby,G. A.; Wharton, C. W.; Thorneley, R. N. F. J. Am.Chem. Soc. 1997, 119,6450-6451] provide strong mutualsupport.