Mechanistic enzymology of CO dehydrogenase from Clostridium thermoaceticum
The final steps in acetyl-CoA biosynthesis by anaerobic bacteria are performed by carbon monoxide dehydrogenase (CODH), a nickel/iron-sulfur protein. An important achievement was to establish conditions under which acetyl-CoA synthesis by purified enzymes equals the in vivo rate of acetate synthesis. Under these optimized conditions we established that the rate limiting step in the synthesis of acetyl-CoA from methyl-H[sub 4]folate, CO and CoA is likely to be the methylation of CODH by the methylated corrinoid/iron-sulfur protein. We then focused on stopped flow studies of this rate limiting transmethylation reaction and established its mechanism. We have studied the carbonylation of CODH by infrared and resonance Raman spectroscopy and determined that the [Ni-Fe[sup 3-4]S[sub 4]]-CO species which has been characterized by magnetic resonance methods can be described as [Ni-X-Fe[sub 3-4]S[sub 4]]-C[equivalent to]O. We showed that this species is the catalytically competent precursor of the carbonyl group of acetyl-CoA. We have made progress in the synthesis of seleno-coenzyme A, which we will use to probe the binding of CoA to CODH. We also have compared the CODH from Methanosarcina thermophila with the C. thermoaceticum enzyme by EPR and electrochemical methods and found that the metal sites of these enzymes are remarkably similar given the evolutionary separation between archaea and bacteria domains.
- Research Organization:
- Nebraska Univ., Lincoln, NE (United States)
- Sponsoring Organization:
- USDOE; USDOE, Washington, DC (United States)
- DOE Contract Number:
- FG02-91ER20053
- OSTI ID:
- 7024909
- Report Number(s):
- DOE/ER/20053-1; ON: DE93002236
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
CARBON MONOXIDE
BIOCONVERSION
OXIDOREDUCTASES
BIOCHEMICAL REACTION KINETICS
ANAEROBIC CONDITIONS
BIOLOGICAL PATHWAYS
CARBON 13
CLOSTRIDIUM
FOLIC ACID
PROGRESS REPORT
SPECIES DIVERSITY
TRACER TECHNIQUES
AMINO ACIDS
AROMATICS
AZAARENES
BACTERIA
CARBON COMPOUNDS
CARBON ISOTOPES
CARBON OXIDES
CARBOXYLIC ACIDS
CHALCOGENIDES
DOCUMENT TYPES
DRUGS
ENZYMES
EVEN-ODD NUCLEI
HEMATINICS
HEMATOLOGIC AGENTS
HETEROCYCLIC COMPOUNDS
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MICROORGANISMS
NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OXIDES
OXYGEN COMPOUNDS
PROTEINS
PTERIDINES
REACTION KINETICS
STABLE ISOTOPES
VITAMIN B GROUP
VITAMINS
090900* - Biomass Fuels- Processing- (1990-)
550201 - Biochemistry- Tracer Techniques