犬溶菌酶基因的克隆、原核表达及产物活性分析  

Clone of synthetic canine Iysozyme gene and its prokaryotic expression and activity analysis

作　　者：朱忠珂[1] 王建华[1] 汪儆[2] 张春雷[1] 蔡青和 

机构地区：[1]西北农林科技大学动物科技学院,陕西杨凌712100 [2]中国农业科学院畜牧研究所,中国北京100094 [3]华芬饲料酶有限公司,广东肇庆526020

出　　处：《西北农林科技大学学报（自然科学版）》2007年第4期55-59,共5页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家星火计划项目(2001EA780037)。

摘　　要：为了探讨重组犬溶菌酶的活性及临床应用前景,人工合成犬溶菌酶基因,构建其原核重组表达载体pPROHTa-rcLYZ,进行原核表达并对表达产物进行活性分析。结果表明,合成的犬溶菌酶基因全长396 bp,编码132 aa;构建的表达载体pPROHTa-rcLYZ含有犬溶菌基因完整的ORF;重组犬溶菌酶的相对分子质量为16 ku;重组犬溶菌酶约占全菌总蛋白32％,纯化后占全菌可溶性蛋白的95％;重组犬溶菌酶最适pH值为6．2,最适温度为38℃,对溶壁微球菌、大肠杆菌、金黄色葡萄球菌和沙门氏菌有较强的溶菌活力,对铜绿假单胞菌、链球菌、枯草杆菌等有一定的抑制作用,对白色念珠菌无抑制作用,对人工感染的犬大肠杆菌病有治疗作用。表明本试验获得了有活性的、具有一定潜在应用价值的犬溶菌酶。In order to investigate the activity of canine lysozyme gene and its prospect in clinical practice,the synthesis method was applied to obtain the complete CDS of canine lysozyme （cLYZ） gene （396 bp, 132 aa） ,and the expression vector pPROHTa-rcLYZ was constructed to study the expression of rcLYZ in E. coli and analyze activity of the recombinant protein on this basis. The results showed that the recombinant expression vector pPROHTa-rcLYZ was constructed successfully, and it contained the entire ORF of canine lysozyme gene;the relative quality of recombinant canine lysozyme is 16 ku. The level of expression and purified protein were 32% and 95 % of the total protein of Escherichia coli, respectively. The optional pH and temperature of the recombinant bacterial strains were 6.2 and 38 ℃. The lysozyme activities analyses revealed that： rcLYZ showed powerful lysozyme activity on some bacterial,e, g. Escherichia coli. , icroccusly sodeikticus, and had restriction activity on some bacterials. Moreover, it was against some bacterial. The recombinant protein contributed to healing of artificial infection of canine E. coli disease. Synthetical lysozyme and expression product had curative effect on dog coli infection. So it can be concluded that prokaryotic canine lysozyme obtained in this experiment had potential application value.

关 键 词：犬溶菌酶基因 犬溶菌酶 原核表达 

分 类 号：Q786[生物学—分子生物学]