Vetter R, Dai J, Mesaeli N, Panagia V, Dhalla N S
Division of Cardiovascular Sciences St. Boniface General Hospital Research Centre, Winnipeg, Canada.
Mol Cell Biochem. 1991 Apr 24;103(1):85-96. doi: 10.1007/BF00229596.
The effect of reagents that modify sulfur-containing amino acid residues in the phosphatidylethanolamine N-methyltransferase was studied in the isolated rat cardiac sarcolemma by employing S-adenosyl-L-[methyl-3H]methionine as a methyl donor. Dithiothreitol protected the sulfhydryl groups in the membrane and caused a concentration- and time-dependent increase of phospholipid N-methylation at three different catalytic sites. This stimulation was highest (9-fold) in the presence of 1 mM MgCl2 and 0.1 microM S-adenosyl-L-[methyl-3H]methionine at pH 8.0 (catalytic site I), and was associated with an enhancement of Vmax without changes in Km for the methyl donor. Thiol glutathione was less stimulatory than dithiothreitol; glutathione disulfide inhibited the phosphatidylethanolamine N-methylation by 50%. The alkylating reagents, N-ethylmaleimide and methylmethanethiosulfonate, inhibited the N-methylation with IC50 of 6.9 and 14.1 microM, respectively; this inhibition was prevented by 1 mM dithiothreitol. These results indicate a critical role of sulfhydryl groups for the activity of the cardiac sarcolemmal phosphatidylethanolamine N-methyltransferase and suggest that this enzyme system in cardiac sarcolemma may be controlled by the glutathione/glutathione disulfide redox state in the cell.
通过使用S-腺苷-L-[甲基-³H]甲硫氨酸作为甲基供体，在分离的大鼠心肌肌膜中研究了修饰磷脂酰乙醇胺N-甲基转移酶中含硫氨基酸残基的试剂的作用。二硫苏糖醇保护膜中的巯基，并在三个不同的催化位点引起磷脂N-甲基化的浓度和时间依赖性增加。在pH 8.0（催化位点I）存在1 mM MgCl₂和0.1 μM S-腺苷-L-[甲基-³H]甲硫氨酸的情况下，这种刺激作用最高（9倍），并且与Vmax的增加相关，而甲基供体的Km没有变化。硫醇型谷胱甘肽的刺激作用比二硫苏糖醇小；谷胱甘肽二硫化物抑制磷脂酰乙醇胺N-甲基化达50%。烷基化试剂N-乙基马来酰亚胺和甲硫基甲烷磺酸盐分别以6.9和14.1 μM的IC50抑制N-甲基化；1 mM二硫苏糖醇可防止这种抑制作用。这些结果表明巯基对心肌肌膜磷脂酰乙醇胺N-甲基转移酶的活性起关键作用，并表明心肌肌膜中的这种酶系统可能受细胞内谷胱甘肽/谷胱甘肽二硫化物氧化还原状态的控制。
