狗母鱼肾脏中三甲胺 - N - 氧化物脱甲基酶的部分纯化及特性研究

Partial purification and characterization of trimethylamine-N-oxide demethylase from lizardfish kidney.

作者信息

Benjakul Soottawat, Visessanguan Wonnop, Tanaka Munehiko

机构信息

Department of Food Technology, Faculty of Agro-Industry, Prince of Songkla University, Hat Yai, Songkhla, 90112, Thailand.

出版信息

Comp Biochem Physiol B Biochem Mol Biol. 2003 Jun;135(2):359-71. doi: 10.1016/s1096-4959(03)00082-4.

DOI:10.1016/s1096-4959(03)00082-4

PMID:12798945

Abstract

Trimethylamine-N-oxide demethylase (TMAOase) from lizardfish (Saurida micropectoralis) was partially purified by acidification and diethylaminoethyl (DEAE)-cellulose chromatography. The enzyme was purified 82-fold with a yield of 65.4%. The optimum pH and temperature were 7.0 and 50 degrees C, respectively. TMAOase was stable to heat treatment up to 50 degrees C and the activation energy was calculated to be 30.5 kJ mol(-1) K(-1). Combined cofactors (FeCl(2), ascorbate and cysteine) were required for full activation. FeCl(2) exhibited a higher stimulating effect on TMAOase activity than FeCl(3). At concentration less than 2 mM, ascorbate was more stimulatory to the activity than cysteine. The activity was tolerant of NaCl concentration up to 0.5 M. The enzyme had a K(m) for TMAO of 16.2 mM and V(max) of 0.35 micromol min(-1) and was able to convert TMAO to dimethylamine (DMA) and formaldehyde. The molecular mass of enzyme was estimated to be 128 kDa based on activity staining.

摘要

通过酸化和二乙氨基乙基（DEAE）-纤维素色谱法对长蛇鲻（Saurida micropectoralis）中的三甲胺 - N - 氧化物脱甲基酶（TMAOase）进行了部分纯化。该酶纯化了82倍，产率为65.4%。最适pH和温度分别为7.0和50℃。TMAOase在高达50℃的热处理下稳定，计算出的活化能为30.5 kJ mol(-1) K(-1)。完全活化需要联合辅因子（FeCl(2)、抗坏血酸和半胱氨酸）。FeCl(2)对TMAOase活性的刺激作用高于FeCl(3)。在浓度低于2 mM时，抗坏血酸对活性的刺激作用比半胱氨酸更强。该酶对高达0.5 M的NaCl浓度具有耐受性。该酶对TMAO的K(m)为16.2 mM，V(max)为0.35 μmol min(-1)，能够将TMAO转化为二甲胺（DMA）和甲醛。基于活性染色，该酶的分子量估计为128 kDa。