CHARACTERIZATION OF IMT1, MYOINOSITOL O-METHYLTRANSFERASE, FROM MESEMBRYANTHEMUM-CRYSTALLINUM

A full-length transcript, Imt1, encoding myo-inositol O-methyltransfer ase (EC 2.1.1.X) from the halophyte Mesembryanthemum crystallinum was expressed in Escherichia coli. The enzyme, IMT1, uses S-adenosyl-L-met hionine to methylate myo-inositol to form D-ononitol. IMT1 with a mono meric mass of 41,000 was isolated by ammonium sulfate fractionation, g el filtration and ion exchange chromatography to apparent purity on so dium dodecyl sulfate-polyacrylamide gel electrophoresis. The N-termina l amino acid sequence of the purified recombinant enzyme was identical to that encoded by the cDNA sequence. The apparent K-m for S-adenosyl methionine was 0.18 mM with a V-max of 1550 pkat/mg protein. The K-m f or myo-inositol was 1.32 mM. The reaction became substrate-inhibited b y concentrations of S-adenosylmethionine greater than 0.5 mM. Inositol methyltransferase was competitively inhibited 50% with 0.01 mM S-aden osyl-homocysteine, while 1 mM homocysteine, homoserine, or adenosine d id not inhibit. The enzyme exhibited a pH optimum of 7.8 and a tempera ture optimum of 37 degrees C. Activity of the isolated inositol methyl transferase was stable when stored at 4 degrees C. (C) 1995 Academic P ress, Inc.