竹叶青蛇毒L-氨基酸氧化酶的纯化、诱导细胞凋亡和抗菌作用  

Purification,Characterization of a Novel Amino-acid Oxidase from Trimeresurus stejneger Venom

作　　者：张玉洁[1] 陈克平[1] 李文辉[2] 张云[2] 

机构地区：[1]江苏大学生命科学院,镇江212013 [2]中国科学院昆明动物研究所,昆明650023

出　　处：《天然产物研究与开发》2006年第1期33-37,共5页Natural Product Research and Development

基　　金：国家自然科学基金项目(30170195)

摘　　要：经过分子筛层析和离子交换层析,我们从竹叶青(Trimeresurus stejnegeri)蛇毒中纯化得到了竹叶青蛇毒L-氨基酸氧化酶(TSV-LAO)。实验表明,TSV-LAO是一种糖蛋白,其分子由非共价连接的两个相同的亚基组成,SDS-聚丙烯酰氨凝胶电泳一个亚基的表观分子量为58 kDa。TSV-LAO酶比活力为1100 U/mg,其分子脱掉糖基化成分后不影响酶活力。TSV-LAO在浓度为1.0μg/mL以上时可以诱导C8166细胞凋亡。TSV-LAO对实验病原微生物具有选择性抗生作用并具有明显的量效关系,对白色念珠菌(ATCC2002)、金黄色葡萄球菌(ATCC2592)和短小芽孢杆菌(CMCCB11207)的最小抑菌浓度分别是0.3,0.4和1.0μg/mL,即使在最高实验浓度10μg/mL,TSV-LAO对其它实验菌株也未显示抑菌作用。A novel L-amino acid oxidase from venom of Chinese snake Trimeresurys stejneger, named TSV-LAO,was purified to homogeneity by two steps of isolation including Sephadex G-100 gel fihration and Q-Sepharose ion-exchange chromatography. TSV-LAO is a glycoprotein and composed of two non-covalent identical subunits with a molecular weight of 58 kDa by SDS-polyacrylamide gel electrophoresis. The specific enzymatic activity of TSV-LAO purified preparations was 1100 U/mg. The enzymatic activity was not lost after deglycosylation. TSV-LAO above 1.0 μg/mL induces apoptosis on C8166 cells dose-dependently. TSV-LAO had antimicrobial activity in a dose-dependent and strain-selective manner. Minimal protein concentration required for total inhibition of the growth of sensitive detected strains of microbe in liquid medium （MIC） was 0.3,0.4 and 1.0 （μ/ mL） on C. albicans （ATCC2002）, S. aureus （ATCC2592） and B. megaterium （CMCCBll207） separately.

关 键 词：蛇毒 L-氨基酸氧化酶 纯化 生化性质 细胞凋亡 抗生作用 

分 类 号：Q554.4[生物学—生物化学]