Potentiometric titrations of carbon monoxide dehydrogenase and properties of the Ni-labile and nonlabile forms of the acetyl-COA synthase active site /
Carbon monoxide dehydrogenase (CODH) from Clostridium thermoaceticum is a bifunctional metalloenzyme, catalyzing the synthesis of acetyl-coenzyme A and reversible oxidation of CO to C02 at two novel Ni-Fe-S active sites (the A and C S,]2111, clusters, respectively). CODH also contains an [Fe, cube,...
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| Format: | Thesis Book |
| Language: | English |
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[Place of publication not identified] :
[publisher not identified] ;
1998.
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| Subjects: | |
| Online Access: | http://proxy.library.tamu.edu/login?url=http://proquest.umi.com/pqdweb?did=737708951&sid=1&Fmt=2&clientId=2945&RQT=309&VName=PQD |
| Summary: | Carbon monoxide dehydrogenase (CODH) from Clostridium thermoaceticum is a bifunctional metalloenzyme, catalyzing the synthesis of acetyl-coenzyme A and reversible oxidation of CO to C02 at two novel Ni-Fe-S active sites (the A and C S,]2111, clusters, respectively). CODH also contains an [Fe, cube, known as the B- cluster. Midpoint potentials of these clusters were obtained by titrating CODH under equilibrium conditions using various partial pressures -of CO in Ar and C02 atmospheres, and simulating EPR signal intensities as a function of potential. Simulations assuming n (#e- involved in reduction) values larger than expected for the individual reactions generally fit better than those which assumed the expected n, indicating positive redox cooperativity. The presence of CO, raised the potentials of the A, B and C clusters, and appeared to increase the strength of CO binding to the reduced A-cluster. C02 appears to stabilize an intermediate EPR-silent state of the C-cluster, and alter the saturation/relaxation properties of the reduced B-cluster. C02 did not inhibit the removal of the labile Ni, but it did inhibit the CO/acetyl-coenzyme A exchange activity. These effects of C02 indicate a significant C02-dependent conformational change affecting the properties of all three clusters in both subunits. Since the enzyme operates in a C02 environment in vivo, the C02-induced conformation may be mechanistically significant. Populations of the (x subunit contain two major forms of A-clusters, a catalytically active form called Ni-Labile and an inactive form called Nonlabile. The Ni component of the Ni-labile form could be reduced either by CO and a catalytic amount of native enzyme or by electrochemically-reduced triquat in the presence of CO. CObinding raised E0(Ni2+/1+),,,, rendering CO and triquat effective reductants. Dithionite did not reduce the Ni-labile form, though its addition to CO/CODH-reduced Ni-labile clusters caused an intracluster electron transfer from the Ni" to the [Fe,S ] 21 cluster. Dithionite reduced the [Fe,S4 ]21 component of the Nonlabile form as well as the cluster of the Ni-labile form once Ni was removed. Ni may not be bridged to the cube in the Nonlabile form. The [Fe,S,]" may transfer electrons to-and-from the redox-active D site during reductive activation, and when bridged to the Ni of the A-cluster, it may modulate the redox and CO-binding properties of the Ni. |
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| Item Description: | Vita. "Major Subject: Chemistry". |
| Physical Description: | xv, 110 leaves : illustrations ; 28 cm. Issued also on microfiche from University Microfilms Inc. |
| Bibliography: | Includes bibliographical references: pages 105-109. |