Acetate-dependent methylation of two corrinoid proteins in extracts of Methanosarcina barkeri

Journal Article · Sun Sep 01 00:00:00 EDT 1991 · Journal of Bacteriology; (United States)

OSTI ID:5755087

Cao, Xianjun; Krzycki, J A ^[1]

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1. Ohio State Univ., Columbus, OH (United States)

Corrinoid proteins have been implicated as methyl carriers in methane formation from acetate, yet specific corrinoid proteins methylated by acetate-derived intermediates have not been identified. In the presence of ATP, H{sub 2}, and bromoethanesulfonic acid, label from {sup 3}H- or 2-{sup 14}C-labeled acetate was incorporated into the protein fraction of cell extracts of Methanosarcina barkeri. Incorporated label was susceptible to photolysis, yielding labeled methane as the anaerobic photolysis product. size exclusion high-pressure liquid chromatography (HPLC) demonstrated the presence of at least three labeled proteins with native molecular sizes of 480, 200, and 29 kDa, while electrophoresis indicated that four major labeled proteins were present. Dual-label experiments demonstrated that these four proteins were methylated rather than acetylated. Two of the proteins (480 and 29 kDa) contained the majority of radiolabel and were stably methylated. After labeling with (2-{sup 14}C)acetate, the stable {sup 14}CH{sub 3}-proteins were partially purified, and {sup 14}CH{sub 3}-cofactors were isolated from each protein. UV-visible spectroscopy and HPLC demonstrated these to be methylated corrinoids. When the protein. UV-visible spectroscopy and HPLC demonstrated these to be methylated corrinoids. When the 480-kDa corrinoid protein was purified to 70% homogeneity, the preparation was found to have subunits of 40 and 30 kDa. The 480-kDa protein but not the 29-kDa protein was methylated during in vitro methanogenesis from acetate and demethylated as methanogenesis ceased, consistent with the involvement of this protein in methane formation.

• OSTI ID: 5755087
• Journal Information: Journal of Bacteriology; (United States), Vol. 173:17; ISSN 0021-9193
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
METHANOGENIC BACTERIA
BIOLOGICAL PATHWAYS
PROTEINS
METHYLATION
ACETATES
BIOSYNTHESIS
CARBON 14 COMPOUNDS
DOUBLE LABELLING
METHANE
TRITIUM COMPOUNDS
ALKANES
BACTERIA
CARBON COMPOUNDS
CARBOXYLIC ACID SALTS
CHEMICAL REACTIONS
HYDROCARBONS
HYDROGEN COMPOUNDS
LABELLED COMPOUNDS
LABELLING
MICROORGANISMS
ORGANIC COMPOUNDS
SYNTHESIS
550201* - Biochemistry- Tracer Techniques