一种来自樟芝的高度稳定的形成层状超氧化物歧化酶：在酵母中的表达及酶学性质

A highly stable cambialistic-superoxide dismutase from Antrodia camphorata: expression in yeast and enzyme properties.

作者信息

Liau Yi-Jen, Wen Lisa, Shaw Jei-Fu, Lin Chi-Tsai

机构信息

Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan.

出版信息

J Biotechnol. 2007 Aug 1;131(1):84-91. doi: 10.1016/j.jbiotec.2007.05.021. Epub 2007 May 31.

DOI:10.1016/j.jbiotec.2007.05.021

PMID:17604867

Abstract

A cDNA encoding a putative superoxide dismutase (SOD) was identified in expressed sequence tags of Antrodia camphorata, a medicinal mushroom found only in Taiwan. The deduced protein was aligned with Mn-SODs and Fe-SODs from other organisms, this SOD showed greater homology to Mn-SOD. Functional A. camphorata SOD protein was overexpressed in yeast and purified. The purified enzyme showed two active forms on a 12.5% native PAGE, a dimer and a monomer. The dimeric protein's half-life of deactivation at 80 degrees C was 7 min, and its thermal inactivation rate constant K(d) was 9.87 x 10(-2)min(-1). The enzyme was stable in a broad pH range from 5-11; in the presence of 0.4M imidazole and 2% SDS. The atomic absorption spectrometric assay showed that 1.0 atom of manganese/iron (9:1) was present in each SOD subunit. The high stability of the enzyme make it better suited than other cambialistic-SODs for use in cosmetics. The SOD also documents its future utility in developing anti-inflammatory agent and in the treatment of chronic diseases.

摘要

在仅发现于台湾的药用真菌樟芝的表达序列标签中，鉴定出一种编码假定超氧化物歧化酶（SOD）的cDNA。将推导的蛋白质与来自其他生物的锰超氧化物歧化酶（Mn-SOD）和铁超氧化物歧化酶（Fe-SOD）进行比对，该SOD与Mn-SOD具有更高的同源性。功能性樟芝SOD蛋白在酵母中过表达并纯化。纯化后的酶在12.5%的非变性聚丙烯酰胺凝胶电泳（native PAGE）上显示出两种活性形式，即二聚体和单体。二聚体蛋白在80℃下的失活半衰期为7分钟，其热失活速率常数K(d)为9.87×10(-2)分钟(-1)。该酶在5至11的广泛pH范围内稳定；在存在0.4M咪唑和2%十二烷基硫酸钠（SDS）的情况下也稳定。原子吸收光谱测定表明，每个SOD亚基中存在1.0个锰/铁原子（9:1）。该酶的高稳定性使其比其他兼性SOD更适合用于化妆品。该SOD还证明了其在开发抗炎剂和治疗慢性疾病方面的未来用途。