新肺癌相关基因hERGIC3的克隆与生物信息学分析

目的构建hERGIC3基因的原核表达载体,并对其进行生物信息学分析,以期全面地了解hERGIC3蛋白的生物学功能。方法采用实时半定量聚合酶链反应（RT-PCR）方法获取人类hERGIC3基因的开放阅读框（ORF）框DNA序列插入pGEM-T Easy载体;选择多个软件对hERGIC3蛋白进行生物信息学分析。结果酶切结果显示,插入序列为目标序列,成功克隆了hERGIC3基因;生物信息学分析结果显示：hERGIC3蛋白由383个氨基酸残基组成,相对分子质量为43.2×10^3,理论等电点为5.68,蛋白比较稳定;hERGIC3为跨膜蛋白,跨膜区为20～42和345～367,膜外区域为1～19和368～383,膜内区域为43～344;二级结构含有110个α螺旋、94条延伸链、184个随机卷曲、15个潜在的磷酸化位点,hERGIC3可能参与了氨基酸、辅酶的生物合成以及脂肪、能量代谢和蛋白质翻译、转运等功能;hERGIC3含有ERGIC_N和COPⅡcoated_ERV 2个蛋白保守结构域,hERGIC3可能与PPKCSH、ERGIC1、ERGIC2、COPA、PSMD11等蛋白有相互作用。结论成功克隆了hERGIC3基因;深入地分析了hERGIC3的结构与功能,为下一步研究hERGIC3基因在肺癌中的病理生理功能奠定了理论基础。

Bioinformatics analysis and clone of hERGIC3 gene related with newly-diagnosed lung cancer

Objective To construct prokaryotic expression vector of hERGIC3 gene and analyze it by bioinformatics,so as to fully understand the biological function of hERGIC3 protein. Methods The pGEM-T Easy vector was inserted into the DNA fragment of open reading frame（ORF） sequence of gene hERGIC3 by real time-polymerase chain reaction（RT-PCR） method;multiple softwares were selected to analyze the hERGIC3 protein by bioinformatics. Results The enzyme digestion showed that the insertion sequence as the targeted sequence cloned the hERGIC3 gene successfully;the bioinformatics analysis showed that hERGIC3 protein was consisted of 383 amino acid residues with the relative molecular mass of 43.2 ×10^3and theoretical isoelectric point of 5.68,and the protein was stable;the hERGIC3 was a transmembrane protein containing transmembrane domains（20-42,345-367）,external membrane areas（1-19,368-383） and internal membrane area（43-344）;the secondary structure was composed with 110 α-helixes,94 extended strands,184 random coils and 15 potential phosphorylation sites,and the hERGIC3 might be involved in biosynthesis of amino acid and coenzyme,energy metabolism of fat,translation and transport of protein;the hERGIC3 contained two conserved domain of protein structure（ERGIC_N and COPⅡcoated_ERV）,and the hERGIC3 might be interacted with PPKCSH,ERGIC1,ERGIC2,COPA,PSMD11 and so on. Conclusion The hERGIC3 gene is cloned successfully;in-depth analysis of the structure and function of hERGIC3 provides theoretical basis for further research of physiological function of hER-GIC3 gene in lung cancer.