34] S-Adenosylmethionine:Indolepyruvate 3-methyltransferase

Abstract

Publisher Summary This chapter discuses the assay method and purification of S-adenosylmethionine. The enzyme catalyzes the transfer of the methyl group from [methyl- 14 C] S-adenosylmethionine to the 3-position of the aliphatic side chain of indolepyruvate. The radioactive reaction product can be extracted from the acidified reaction mixture with butyl acetate and then counted in a liquid scintillation spectrometer. In purification procedure, all steps are performed at 0–4° unless otherwise stated. The first step is growth of cultures and preparation of the crude cell-free extract. Further procedure involves ammonium sulfate treatment, sephadex chromatography, diethylaminoethyl (DEAE)-sephadex chromatography, bio-gel A-5m chromatography. By this procedure, the enzyme may be purified approximately 110-fold with an overall yield of 40–45%. The enzyme can be stored at 2° for at least 3 weeks without significant loss of activity.