title: Purification and characterization of homodimeric methylmalonyl-CoA mutase from Sinorhizobium meliloti.
abstract: High activity (>60 munit/mg protein) of 5'-deoxyadenosylcobalamin-dependent methylmalonyl-CoA mutase (EC 5.4.99.2) was constantly found during growth of a strain of the root-nodule-forming bacterium Sinorhizobium meliloti harboring an extra plasmid-encoded copy of the methylmalonyl-CoA-mutase-encoding bhbA gene. The enzyme was purified to homogeneity and characterized. The purified enzyme was found to be a colorless apo-form. The apparent molecular weight of the enzyme was calculated to be 165,000+/-5,000 by Superdex 200 HR gel filtration. SDS-PAGE of the purified enzyme resolved one protein band with an apparent molecular mass of 80.0+/-2.0 kDa, indicating that the S. meliloti enzyme is composed of two identical subunits. The NH(2)-terminal sequence was identical to that predicted from the bhbA nucleotide sequence. Monovalent cations were required for enzyme activity.
authors.affiliation: Department of Health Science, Kochi Women's University, 780-8515, Kochi, Japan.
authors.forename: Emi
authors.initials: E
authors.lastname: Miyamoto
authors.forename: Fumio
authors.initials: F
authors.lastname: Watanabe
authors.forename: Trevor C
authors.initials: TC
authors.lastname: Charles
authors.forename: Ryoichi
authors.initials: R
authors.lastname: Yamaji
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Inui
authors.forename: Yoshihisa
authors.initials: Y
authors.lastname: Nakano
mesh_terms: D001051:Apoenzymes / Q000302:isolation & purification; D002850:Chromatography, Gel; D003038:Cobamides; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D020035:Holoenzymes / Q000378:metabolism; D007700:Kinetics; D008765:Methylmalonyl-CoA Mutase / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D021122:Protein Subunits; D016962:Sinorhizobium meliloti / Q000201:enzymology* / Q000254:growth & development
chemical_list: D001051:Apoenzymes; D003038:Cobamides; D020035:Holoenzymes; D021122:Protein Subunits; D008765:Methylmalonyl-CoA Mutase; C000913:cobamamide