Binding of S-adenosylhomocysteine to hydroxyindole O-methyltransferase.
Biochim Biophys Acta 1984 May 31;787(1 ) 1-7 (S-adenosyl-L-methionine:N-acetylserotonin O-methyltransferase, EC 2.1.1.4) purified from bovine pineal gland forms a complex with S-adenosylhomocysteine, one of the products of the reaction catalyzed by the enzyme. The binding of S-adenosylhomocysteine to the enzyme has been characterized by the use of S-[8-14C]adenosylhomocysteine or S-[U -14C]adenosylhomocysteine. The complex did not dissociate during filtration through Sephadex G-25. Long-term dialysis against ligand-free phosphate buffer did bot dissociate the bound S-adenosylhomocysteine. S-Adenosylhomocysteine co-migrated with hydroxyindole O-methyltransferase on disc polyacrylamide gel electrophoresis. Although the complex was stable even under nonequilibrium conditions, the bound S- adenosylhomocysteine was separated into adenosine and homocysteine in the presence of S- adenosylhomocysteine hydrolase. The binding of S-adenosylhomocysteine was optimal at pH 7.0, but was not dependent on temperature. Scatchard plots showed that Kd was 6.5 X 10(-9) M and the maximal binding was 1 mol of S-adenosylhomocysteine per subunit of the enzyme. Hydroxyindole O-methyltransferase cannot form a stable complex with S-adenosylmethionine, and the addition of excess amounts of S-adenosylmethionine impairs the binding of S-adenosylhomocysteine to the enzyme. The product inhibition by S-adenosylhomocysteine may be based on the binding of S-adenosylhomocysteine to the enzyme with high affinity and on the stable enzyme-product complex formation during the transmethylation reaction.
D000121:Acetylserotonin O-Methyltransferase / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D006710:Homocysteine / Q000031:analogs & derivatives*; D007700:Kinetics; D008780:Methyltransferases / Q000378:metabolism*; D010870:Pineal Gland / Q000201:enzymology*; D011485:Protein Binding; D012435:S-Adenosylhomocysteine / Q000378:metabolism*
D006710:Homocysteine; D012435:S-Adenosylhomocysteine; D008780:Methyltransferases; D000121:Acetylserotonin O-Methyltransferase