S-Adenosylmethionine synthetase from Escherichia coli.
Adenosylmethionine (AdoMet) synthetase has been purified to homogeneity from Escherichia coli. For this purification, a strain of E. coli which was derepressed for AdoMet synthetase and which harbors a plasmid containing the structural gene for AdoMet synthetase was constructed. This strain produces 80-fold more AdoMet synthetase than a wild type E. coli. AdoMet synthetase has a molecular weight of 180,000 and is composed of four identical subunits. In addition to the synthetase reaction, the purified enzyme catalyzes a tripolyphosphatase reaction that is stimulated by AdoMet. Both enzymatic activities require a divalent metal ion and are markedly stimulated by certain monovalent cations. AdoMet synthesis also takes place if adenyl-5'yl imidodiphosphate (AMP-PNP) is substituted for ATP. The imidotriphosphate (PPNP) formed is not hydrolyzed, permitting dissociation of AdoMet formation from tripolyphosphate cleavage. An enzyme complex is formed which contains one equivalent (per subunit) of adenosylmethionine, monovalent cation, imidotriphosphate, and presumably divalent cation(s). The rate of product dissociation from this complex is 3 orders of magnitude slower than the rate of AdoMet formation from ATP. Studies with the phosphorothioate derivatives of ATP (ATP alpha S and ATP beta S) in the presence of Mg2+, Mn2+, or Co2+ indicate that a divalent ion is bound to the nucleotide during the reaction and provide information on the stereochemistry of the metal-nucleotide binding site.
D000266:Adenylyl Imidodiphosphate; D002857:Chromium / Q000494:pharmacology; D003035:Cobalt / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008716:Methionine Adenosyltransferase / Q000378:metabolism*; D008970:Molecular Weight; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D013237:Stereoisomerism; D014166:Transferases / Q000378:metabolism*
D002857:Chromium; D000266:Adenylyl Imidodiphosphate; D003035:Cobalt; D008345:Manganese; D014166:Transferases; D008716:Methionine Adenosyltransferase; D010744:Phosphoric Monoester Hydrolases; D008274:Magnesium