摘要
目的分析华支睾吸虫乳酸脱氢酶基因及其编码蛋白的结构和特性,用于指导其生物学功能的实验研究。方法利用多种生物信息在线分析工具和分析软件包,从华支睾吸虫全长cDNA质粒文库的表达序列标签(EST)中识别乳酸脱氢酶(CsLDH)基因,预测该基因编码蛋白质的理化特性、氨基酸修饰位点、功能域、亚细胞定位、拓扑结构、二级结构、三维结构及酶学特性和免疫学特性等。结果该基因全长1224bp,编码区为79～1062,编码328aa,理论分子量为35633Mr,等电点为8.13,半衰期长,理化性质稳定,含有多个潜在的磷酸化和酯酰化位点。预测该蛋白有3个跨膜区,其拓扑结构为N端在膜内侧、C端在膜外侧。有两个主要的线性抗原表位aa10～aa20和aa94～aa102,后者位于膜外,与日本血吸虫LDH相应表位完全一致,与人和小鼠LDH相应表位仅有1个氨基酸差异。该表位中Arg102是酶催化中心的关键氨基酸之一,模拟三维结构显示组成催化中心的另外两个关键氨基酸残基Asp162和His189的空间位置也靠近该表位,酶的催化中心贯穿膜内外。结论推测华支睾吸虫乳酸脱氢酶不仅负责将糖酵解产生的丙酮酸转化为乳酸,而且还能将乳酸直接排出体外;可作为筛选水溶性抗华支睾吸虫药物的靶标;可介导特异性抗体对虫体的ADCC作用和补体的杀伤作用,以及对酶活性的抑制作用,是一个重要的疫苗侯选抗原。
Objective Bioinformatics analyze the structure and characteristics of Clonorchis sinenzis lactate dehydrogenase to guide the experimental research on its biological function and application. Method Utilizing the tools in bioinformaties webs sites such as NCBI（http：//www.nebi.nlm.nih.gov/）, ExPaSy （http：//www.expasy.org/） and other bioinformatics software package such as PCGENE, Vector NTI suite 8.0, to recognize lactate dehydrogenase gene from the expression sequence tags （ESTs） of Clonorchis sinensis full-length eDNA： plasmid libratory, then to predict the physical-chemical characteristics of the putative protein, as well as the structural characteristics such as epitopes, posttranslational modification sites, domains, subeelluar location, topological structure, second structures, and 3D structure, and so on. Result The gene is 1224 bp in length, coding 328 amino acids in the coding region of 79-1062, and is a complete full-length gene compared with the homologues in C, enBank. The protein contains three transmembrane regions and two main linear epitopes, of which epitope aa94-aa102 is located in the loop outside of membrane and contain a key residue Arg162 which is consisting of the active center of the enzyme; inside of membrane, there is substrate pyruvie acid binding site, the catalytic center of the enzyme across the membrane. The most similar homologue is Schistosoma japonicum lactate dehydrogenase,with nearly the same topolologieal structure and 3D conformation, and share an identical outside linear epitopo of aa94-aa102 and in which there is only one residue difference with that of the human and mouse hosts. Conclusion CsLDH is functionally responsible for the convertion of pyruvic acid which is the product of glycolysis to lactate and transport the lactate out of the cell, so it is a promising target for screening water- soluble drugs and vaccine.
出处
《热带医学杂志》
CAS
2007年第12期1145-1148,F0002,共5页
Journal of Tropical Medicine
基金
"863"专题(No.2006AA02Z422)
广东省重大科技专项(No.2004A30801004)。
关键词
华支睾吸虫
乳酸脱氢酶
结构与功能
生物信息学
Clonorchis sinensis
lactate dehydrogenase
structure and function
bioinformatics
