Purification and partial characterization of rat kidney histamine-N-methyltransferase.
Histamine-N-methyltransferase (EC 2.1.1.8) was purified 1700-fold with a yield of 9% from rat kidney. Purification included ammonium sulfate precipitation, linear gradient DEAE-cellulose chromatography and S-adenosylhomocysteine affinity chromatography. The purified enzyme preparation showed a single protein band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecular weight of 35000. The isoelectric point of the enzyme was at pH 5.2. The purified enzyme preparation did not contain detectable amounts of histamine. The purified enzyme was totally inhibited in 100 microM parahydroxymercuric benzoate and in 10 microM iodoacetamide, and it was found to be stabilized with dithiothreitol (1 mM), suggesting that the enzyme has an SH-group in the active center. The Km values for histamine and S-adenosylmethionine were 6.0 and 7.1 microM, respectively. 50% inhibition of histamine-N-methyltransferase was obtained at 28 microM S-adenosylhomocysteine and 100 microM methylhistamine. The purified enzyme was slightly inhibited in 1 mM methylthioadenosine. Histamine in concentrations higher than 25 microM caused substrate inhibition.
D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002848:Chromatography, DEAE-Cellulose; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006637:Histamine N-Methyltransferase / Q000302:isolation & purification* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006902:Hydroxymercuribenzoates / Q000494:pharmacology; D007460:Iodoacetamide / Q000494:pharmacology; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D008297:Male; D008761:Methylhistamines / Q000494:pharmacology; D008780:Methyltransferases / Q000302:isolation & purification*; D051381:Rats; D011919:Rats, Inbred Strains; D012435:S-Adenosylhomocysteine / Q000494:pharmacology; D012436:S-Adenosylmethionine / Q000378:metabolism; D013872:Thionucleosides / Q000494:pharmacology
D006902:Hydroxymercuribenzoates; D008761:Methylhistamines; D013872:Thionucleosides; C030601:4-hydroxymercuribenzoate; C021714:2-methylthioadenosine; D012436:S-Adenosylmethionine; D012435:S-Adenosylhomocysteine; D008780:Methyltransferases; D006637:Histamine N-Methyltransferase; C026259:2-methylhistamine; D000241:Adenosine; D004229:Dithiothreitol; D007460:Iodoacetamide