Enzymological studies of one-carbon reactions in the pathway of acetate utilization by methanogenic bacteria: Annual technical report for the period February 1, 1988--January 31, 1989

Technical Report · Sun Jan 01 04:00:00 EST 1989

OSTI ID:6463189

Ferry, J G

Our current understanding of the pathway of acetate conversion to methane in Methanosarcina thermophila is depicted. Our accomplishments this past year include phosphotransacetylase, the corrinoid/Fe-S protein component of the carbon monoxide dehydrogenase, and ferredoxin were characterized in further detail; two methyl transferase activities were resolved; and putative positive clones containing the phosphotransacetylase gene were isolated from a lambda gt 11 DNA library of M. thermophila. The phosphotransacetylase was purified to electrophoretic homogeneity and characterized. The specific activity was 2555 micromoles of acetyl-CoA/min/mg. Optimum activity is obtained between 35-45 /degree/C and pH 6.5-7.5 in the presence of potassium or ammonium ions(ca. 100 mM). The enzyme is purified as a monomer of Mr 43,000. The Km and Vmax for CoASH and acetyl phosphate are 91 and 165 micromolar, and 5517 and 4025 micromoles/min/mg. Anti-phosphotransacetylase antibodies were raised in rabbits which were used in an immunoblot of crude extracts from acetate- or methanol-grown cells; the results indicate regulation of synthesis in response to the growth substrate. A mixed oligonucleotide probe was synthesized using the DNA sequence deduced from the N-terminal amino acid sequence. 6 figs.

• Research Organization: Virginia Polytechnic Inst. and State Univ., Blacksburg (USA)
• DOE Contract Number: FG05-87ER13730
• OSTI ID: 6463189
• Report Number(s): DOE/ER/13730-2; ON: DE89006907
• Country of Publication: United States
• Language: English

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59 BASIC BIOLOGICAL SCIENCES
ACETATES
ALKANES
BACTERIA
BIOSYNTHESIS
CARBON COMPOUNDS
CARBON MONOXIDE
CARBON OXIDES
CARBOXYLIC ACID SALTS
CATABOLISM
CHALCOGENIDES
CYTOCHROMES
DOCUMENT TYPES
ENZYME ACTIVITY
ENZYMES
FERREDOXIN
FRACTIONATION
HYDROCARBONS
METABOLISM
METALLOPROTEINS
METHANE
METHANOGENIC BACTERIA
MICROORGANISMS
ORGANIC COMPOUNDS
OXIDES
OXIDOREDUCTASES
OXYGEN COMPOUNDS
PIGMENTS
PROGRESS REPORT
PROTEINS
SEPARATION PROCESSES
SYNTHESIS