Isolation of O-demethylase, an ether-cleaving enzyme system of the homoacetogenic strain MC.
The O-demethylase of the methylotrophic homoacetogenic bacterium strain MC was purified to apparent homogeneity. The enzyme system consisted of four different components that were designated A, B, C, and D according to their elution sequence from the anionic-exchange chromatography column. All four components were essentially required for catalysis of the transfer of the methyl group from phenyl methyl ethers to tetrahydrofolate. According to gel filtration and SDS-PAGE, components A and B were monomers with apparent molecular masses of approximately 26 kDa (subunit 25 kDa) and 36 (subunit 41 kDa), respectively; component C appeared to be a trimeric protein (195 kDa, subunit 67 kDa); and component D was probably a dimer (64 kDa, subunit 30 kDa). Component A contained one corrinoid per monomer. In crude extracts, component D appeared to be the rate-limiting protein for the complete methyl transfer reaction. Additional requirements for the reaction were ATP and low-potential reducing equivalents supplied by either titanium(III) citrate or H2 plus hydrogenase purified from strain MC.
Kaufmann
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F
Institut für Mikrobiologie, Universität Stuttgart, Allmandring 31, D-70569 Stuttgart, Germany.
Wohlfarth
G
G
Diekert
G
G
D019342:Acetic Acid / Q000378:metabolism*; D000840:Anisoles / Q000378:metabolism*; D001673:Biodegradation, Environmental; D002245:Carbon Dioxide / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000201:enzymology*; D006859:Hydrogen / Q000378:metabolism; D008031:Lignin / Q000378:metabolism; D008780:Methyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D008956:Models, Chemical; D009097:Multienzyme Complexes / Q000378:metabolism; D010090:Oxidoreductases, O-Demethylating / Q000302:isolation & purification* / Q000378:metabolism; D011487:Protein Conformation; D013053:Spectrophotometry; D013763:Tetrahydrofolates / Q000378:metabolism*
D000840:Anisoles; D009097:Multienzyme Complexes; D013763:Tetrahydrofolates; D002245:Carbon Dioxide; C030371:5,6,7,8-tetrahydrofolic acid; D006859:Hydrogen; D008031:Lignin; C060998:anisole; D010090:Oxidoreductases, O-Demethylating; D008780:Methyltransferases; D019342:Acetic Acid