PURIFICATION AND CHARACTERIZATION OF A NOVEL METHYLTRANSFERASE RESPONSIBLE FOR BIOSYNTHESIS OF HALOMETHANES AND METHANETHIOL IN BRASSICA-OLERACEA
PURIFICATION AND CHARACTERIZATION OF A NOVEL METHYLTRANSFERASE RESPONSIBLE FOR BIOSYNTHESIS OF HALOMETHANES AND METHANETHIOL IN BRASSICA-OLERACEA
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DOI：
10.1074/jbc.270.16.9250
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发表时间：
1995-04-21
影响因子：
4.8
通讯作者：
SAINI, HS
中科院分区：
文献类型：
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作者：
ATTIEH, JM;HANSON, AD;SAINI, HS
一种新的s -腺苷- l-蛋氨酸：卤化物/二硫化物甲基转移酶（EC 2.1.1）。该酶催化S腺苷基- l-蛋氨酸依赖的卤化物碘化物、溴化物和氯化物甲基化为单卤甲烷，二硫化物甲基化为甲硫醇。该酶的双重功能通过对卤化物和亚硫化物甲基化活性的相同比例的共纯化，以及对碘化物和亚硫化物替代底物之间竞争的研究得到了证明。纯化过程包括凝胶过滤、阴离子交换层析和腺苷琼脂糖亲和层析。用染色质聚焦柱洗脱该蛋白，pI值为4.8。卤化物甲基化的最适pH值（5.5 ~ 7.0）与二硫化物甲基化的最适pH值（7.0 ~ 8.0）不同。天然蛋白和变性蛋白的分子质量值分别为29.5和28 kDa，表明该酶为单体，对碘化物的特异性常数最高，对二硫化物的特异性常数次之，底物相互作用动力学和产物抑制模式符合有序毕比机制。
A novel S-adenosyl-L-methionine:halide/bisulfide methyltransferase (EC 2.1.1.-) was purified approximately 1000-fold to apparent homogeneity from leaves of Brassica oleracea, The enzyme catalyzed the S adenosyl-L-methionine-dependent methylation of the halides iodide, bromide, and chloride to monohalomethanes and of bisulfide to methanethiol. The dual function of the enzyme was demonstrated through co-purification of the halide- and bisulfide-methylating activities in the same ratio and by studies of competition between the alternative substrates iodide and bisulfide, The purification procedure included gel filtration, anion exchange chromatography, and affinity chromatography on adenosine-agarose. Elution of the protein from a chromate-focusing column indicated a pI value of 4.8. The pH optimum of halide methylation (5.5-7.0) was different from that of bisulfide methylation (7.0-8.0). The molecular mass values for the native and denatured protein were 29.5 and 28 kDa, respectively, suggesting that the active enzyme is a monomer, The enzyme had the highest specificity constant for iodide and the next highest for bisulfide, Substrate interaction kinetics and product inhibition patterns were consistent with an Ordered Bi Bi mechanism.