An EPR and electron nuclear double resonance investigation of carbon monoxide binding to hydrogenase I (bidirectional) from Clostridium pasteurianum W5.
1986; Elsevier BV; Volume: 261; Issue: 29 Linguagem: Inglês
10.1016/s0021-9258(18)67051-6
ISSN1083-351X
AutoresJoshua Telser, Michael J. Benecky, Michael W. W. Adams, Leonard E. Mortenson, Brian M. Hoffman,
Tópico(s)Hydrogen Storage and Materials
ResumoPrevious Mossbauer and electron nuclear double resonance (ENDOR) studies of oxidized hydrogenase I (bidirectional) from Clostridium pasteurianum W5 demonstrated that this enzyme contains two diamagnetic [4Fe-4SJ2+ clusters and an iron-sulfur center of unknown structure and composition that is characterized by its novel Miissbauer and ENDOR properties.In the present study we combine ENDOR and EPR measurements to show that the novel cluster contains 3-4 iron atoms.In addition, we have used EPR and ENDOR spectroscopies to investigate the effect of binding the competitive inhibitor carbon monoxide to oxidized hydrogenase I, using "C-labeled CO and enzyme isotopically enriched in "7Fe.Treatment of oxidized enzyme with CO causes theg-tensor of the paramagnetic center to change from rhombic to axial symmetry.The observation of a "C signal by ENDOR spectroscopy and analysis of the EPR broadening show that a single CO covalently binds to the paramagnetic center.The 13C hyperfine coupling constant (Ac = 21 MHz) is within the range observed for inorganic iron-carbonyl clusters.The observation of S7Fe ENDOR signals from two types of iron site (I Alc I -30-34 MHz; I Azc I -6 MHz) and resolved 67Fe hyperfine interactions in the EPR spectrum from two nuclei characterized byI Alc I confirm that the iron-sulfur cluster remains intact upon CO coordination, but show that CO binding greatly changes the "Fe hyperfine coupling constants.Hydrogenases, a class of enzymes (EC 1.12 and 1.18) that catalyze the reaction, 2H+ + 2e-=Hz, have been isolated from a variety of microorganisms.Although these enzymes vary considerably in their molecular composition, metal content, specific activity, and sensitivity to inactivation by oxygen (for review, see Ref. I), all contain iron-sulfur clusters and most of them also contain nickel (2).Two hydrogenases containing iron but not nickel have been isolated from the anaerobic Nzfixing bacterium, Clostridium pasteurianum W5, and one of these is the subject of this report.Referred to as hydrogenase I, it has a molecular weight of 60,000 and contains 12 iron
Referência(s)