52000Ro/SSA抗原结构分析及抗原片段克隆

目的 预测相对分子量为52000的Ro/SSA自身抗原的可能表位,并选择性克隆含亮氨酸拉链氨基酸序列的抗原片段。方法 用蛋白结构分析软件对52000Ro/SSA抗原结构进行分析,从人心脏来源的cDNA中通过聚合酶链反应(PCR)扩增得到编码第119至264位氨基酸片段的cDNA,插入表达载体,转染大肠杆菌并表达融合蛋白。结果 计算机分析显示该抗原中有α螺旋结构,整个分子抗原性强,柔韧性一般,表面可及性差。PCR产物长440bp,重组融合蛋白相对分子质量为28000,测序鉴定保证了研究的正确性。结论 52000Ro/SSA多肽抗原性好,所获得的抗原片段为今后研究亮氨酸拉链氨基酸序列在该抗原表位形成中的作用奠定了基础。

Struture Analysis and Selective Fragment Cloning of 52 kD Ro/SSA Antigen

Objective To detect the possible epitopes of 52 000 dalton Ro/SSA autoantigen and selectively clone the antigen fragment containing the leucine zipper motif. Methods The structure of 52 000 dalton Ro/SSA autoantigen was analyzed by protein structure analysis system. The cDNA encoding the antigen fragments from amino acid (aa) 119 to 264 was amplified from cDNA first strands of human heart by polymerase chain reaction (PCR). The PCR product was inserted into the vector pMTY4, and transfected E. coli pop 2136 for fusion protein production. Results Computer analysis showed that the 52 000 dalton Ro/SSA antigen had alpha helixes in its structure, and had good antigenicity, moderate flexibility and poor surface probability. The PCR product was 440bp in length. The recombinant fusion protein was 28 000 dalton. DNA sequencing proved the accuracy of the research. Conclusion The 52 000 dalton Ro/SSA autoantigen has good antigenicity. The cloning of the antigen fragment will help to elucidate the possible role of leucine zipper motif in forming the epitope of the antigen.