Purification and characterization of S-adenosyl-L-methionine: Benzoic acidcarboxyl methyltransferase, the enzyme responsible for biosynthesis of thevolatile ester methyl benzoate in flowers of Antirrhinum majus

S-Adenosyl-L-methionine:benzoic acid carboxyl methyltransferase (BAMT) cata lyzes the transfer of the methyl group of S-adenosyl-L-methionine (SAM) to the carboxyl group of benzoic acid to make the volatile ester methyl benzoa te, one of the most abundant scent compounds of snapdragon, Antirrhinum ma, ius. The enzyme was purified from upper and lower petal lobes of 5- to 10-d ay-old snapdragon flowers using DE53 anion exchange, Phenyl-Sepharose 6FF, and Mono-Q chromatography, The purified protein has a pH optimum of 7.5 and is highly specific for benzoic acid, with no activity toward several other naturally occurring substrates such as salicylic acid, cinnamic acid, and their derivatives. The molecular mass values for native and denatured prote in were 100 and 49 kDa, respectively, suggesting that the active enzyme is a homodimer, The addition of monovalent cations K+ and NH4+ stimulates BAMT activity by a factor of 2, whereas the addition of Fe2+ and Cu2+ has a str ong inhibitory effect. Plant-purified BAMT has K-m values of 28 mu M and 1. 1 mM for SAM and benzoic acid, respectively (87 mu M and 1.6 mM, respective ly, for plant BAMT expressed in Escherichia coli), product inhibition studi es showed competitive inhibition between SAM and S-adenosyl-L-homocysteine (SAH), with a K-i of 7 mu M, and noncompetitive inhibition between benzoic acid and SAH, with a K-i of 14 mu M. (C) 2000 Academic Press.