Abstract

The objectives of this research are to elucidate the role of 8-hydroxy-7-demethyl-5-deazaflavin coenzymes and nickel-dependent enzymes in the biogenesis of natural gas from C02 and H2 by methanogenic bacteria. The deazaflavins are structural and functional hybrids between nicotinamide and flavin coenzymes and are used as low potential redox currencies. We will elucidate redox stereochemistry, modes of deazaflavin biosynthesis, enzymic nucleotidylation reactions and other roles as coenzymes in the action of DNA photolyase, involved in photomonomerization of thymine dimer lesions in DNA. Over the past five years four novel nickel-containing enzymes have been detected and purified from methanognic bacteria. We are studying two nickel-containing hydrogenases, nickel-linked methyl-S-coenzyme M reductase, and a nickel-dependent CO dehydrogenase as nickel hydrogenation biocatalyst, nickel desulfurization biocatalyst, and nickel carbonylation biocatalyst, respectively. We shall continue efforts to analyze the structure of the nickel active sites to identify ligands nickel oxidation states and geometries to understand catalytic function.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM031574-07
Application #
3279690
Study Section
Biochemistry Study Section (BIO)
Project Start
1988-02-01
Project End
1992-01-31
Budget Start
1989-02-01
Budget End
1990-01-31
Support Year
7
Fiscal Year
1989
Total Cost
Indirect Cost

  Institution

Name
Harvard University
Department
Type
Schools of Medicine
DUNS #
082359691
City
Boston
State
MA
Country
United States
Zip Code
02115

  Publications

SooHoo, C K; Hollocher, T C; Kolodziej, A F et al. (1991) Extended x-ray absorption fine structure and electron paramagnetic resonance of nitrous oxide reductase from Pseudomonas aeruginosa strain P2. J Biol Chem 266:2210-8
Raybuck, S A; Ramer, S E; Abbanat, D R et al. (1991) Demonstration of carbon-carbon bond cleavage of acetyl coenzyme A by using isotopic exchange catalyzed by the CO dehydrogenase complex from acetate-grown Methanosarcina thermophila. J Bacteriol 173:929-32
Zhang, C S; Hollocher, T C; Kolodziej, A F et al. (1991) Electron paramagnetic resonance observations on the cytochrome c-containing nitrous oxide reductase from Wolinella succinogenes. J Biol Chem 266:2199-202
Alex, L A; Reeve, J N; Orme-Johnson, W H et al. (1990) Cloning, sequence determination, and expression of the genes encoding the subunits of the nickel-containing 8-hydroxy-5-deazaflavin reducing hydrogenase from Methanobacterium thermoautotrophicum delta H. Biochemistry 29:7237-44
Ramer, S E; Raybuck, S A; Orme-Johnson, W H et al. (1989) Kinetic characterization of the [3'-32P]coenzyme A/acetyl coenzyme A exchange catalyzed by a three-subunit form of the carbon monoxide dehydrogenase/acetyl-CoA synthase from Clostridium thermoaceticum. Biochemistry 28:4675-80
Reeve, J N; Beckler, G S; Cram, D S et al. (1989) A hydrogenase-linked gene in Methanobacterium thermoautotrophicum strain delta H encodes a polyferredoxin. Proc Natl Acad Sci U S A 86:3031-5
Raybuck, S A; Bastian, N R; Orme-Johnson, W H et al. (1988) Kinetic characterization of the carbon monoxide-acetyl-CoA (carbonyl group) exchange activity of the acetyl-CoA synthesizing CO dehydrogenase from Clostridium thermoaceticum. Biochemistry 27:7698-702
Wackett, L P; Hartwieg, E A; King, J A et al. (1987) Electron microscopy of nickel-containing methanogenic enzymes: methyl reductase and F420-reducing hydrogenase. J Bacteriol 169:718-27
Livingston, D J; Fox, J A; Orme-Johnson, W H et al. (1987) 8-Hydroxy-5-deazaflavin-reducing hydrogenase from Methanobacterium thermoautotrophicum: 2. Kinetic and hydrogen-transfer studies. Biochemistry 26:4228-37
Wackett, L P; Honek, J F; Begley, T P et al. (1987) Substrate analogues as mechanistic probes of methyl-S-coenzyme M reductase. Biochemistry 26:6012-8

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