大鼠肝脏微粒体中二乙基二硫代氨基甲酸的S-甲基化作用

S-methylation of diethyldithiocarbamic acid in rat liver microsomes.

Lill J S, Mays D C, Lipsky J J

Department of Pharmacology, Mayo Medical School, Mayo Clinic/Foundation, Rochester, MN 55905, USA.

Xenobiotica. 1996 Oct;26(10):1025-33. doi: 10.3109/00498259609167420.

Abstract

Human hepatic thiol methyltransferase (TMT) is a microsomal enzyme known to catalyse the in vitro S-methylation of diethyldithiocarbamic acid (DDC) to form diethyldithiocarbamic acid methyl ester (methyl DDC). In vivo data are needed to investigate further the biotransformation of DDC to methyl DDC. Thus, we have characterized the in vitro conversion of DDC to methyl DDC using rat liver microsomes with the ultimate goal of establishing an animal model. 2. Formation of methyl DDC in rat liver microsomes was confirmed by hplc analysis. 3. Rat liver microsomes catalysed methylation of DDC with low and high Km's of 5 +/- 6 and 260 +/- 80 microM respectively and with corresponding Vmax's of 0.09 +/- 0.05 and 0.59 +/- 0.04 nmol/min/mg protein. 4. Rat liver TMT activity was maximally inhibited by 57 +/- 6% by 1000 microM 2,3-dichloro-alpha-methylbenzylamine (DCMB), whereas human TMT was completely inhibited. The concentration of half maximal inhibition of rat TMT for DCMB was 2 microM. 5. Incomplete inhibition of rat TMT activity by DCMB suggests a possible alternative pathway of methylation.

摘要

人肝硫醇甲基转移酶（TMT）是一种微粒体酶，已知可在体外催化二乙氨基二硫代甲酸（DDC）的S-甲基化反应，生成二乙氨基二硫代甲酸甲酯（甲基DDC）。需要体内数据来进一步研究DDC向甲基DDC的生物转化。因此，我们利用大鼠肝微粒体对DDC向甲基DDC的体外转化进行了表征，最终目标是建立一个动物模型。2. 通过高效液相色谱分析证实了大鼠肝微粒体中甲基DDC的形成。3. 大鼠肝微粒体催化DDC的甲基化反应，其低Km值为5±6微摩尔，高Km值为260±80微摩尔，相应的Vmax值分别为0.09±0.05和0.59±0.04纳摩尔/分钟/毫克蛋白质。4. 1000微摩尔的2,3-二氯-α-甲基苄胺（DCMB）可使大鼠肝TMT活性最大程度地抑制57±6%，而人TMT则被完全抑制。DCMB对大鼠TMT的半数最大抑制浓度为2微摩尔。5. DCMB对大鼠TMT活性的不完全抑制表明可能存在甲基化的替代途径。