猪PKM2基因的序列分析与组织表达及亚细胞定位

Sequence analysis and tissue expression and subcellular localization of porcine PKM2 gene

运用RT–PCR技术克隆猪PKM2基因的编码区序列,对该序列进行生物信息学分析,研究PKM2基因的组织表达及亚细胞定位。结果显示:猪PKM2基因CDS全长1 596 bp,编码531个氨基酸;预测其蛋白相对分子质量为5.79×104,等电点为7.96,含有多个磷酸化位点,为稳定蛋白;PKM2蛋白含有丙酮酸激酶家族的barreldomain和alpha/beta domain 2个结构域,其蛋白二级结构中α–螺旋和无规则卷曲占主要类型;猪PKM2蛋白序列在物种间非常保守,系统进化树分析表明,该蛋白与兔的亲缘关系最近;组织表达显示,猪PKM2基因在肾、小肠中相对高表达,而在肝脏与肌肉中相对低表达;亚细胞定位显示,PKM2蛋白在猪3D4/21和PK15细胞中表达于细胞质,而不表达于细胞核。

The coding region of PKM2 was cloned by RT-PCR,and was subjected to sequence bioinformatics analysis,tissue expression profiling and subcellular localization.The results showed that the CDS of porcine PKM2 gene was 1 596 bp in length,which encodes 531 amino acids with the molecular weight 5.79×10^4,the isoelectric point of 7.96,and multiple phosphorylation sites.PKM2 protein had two domains,pyruvate kinase,barrel domain and pyruvate kinase,alpha/beta domain.Regarding to secondary structure,theα-helices and random coil accounted for the main types in the PKM2 protein.The PKM2 protein sequence was very conserved among species,and phylogenetic tree analysis showed that the procine PKM2 protein is closely related to rabbits.The tissue expression profiling revealed that PKM2 gene was highly expressed in kidney and small intestine,and was relatively low expressed in liver and muscle.Subcellular localization showed that the PKM2 protein is expressed in the cytoplasm in porcine 3D4/21 and PK15 cells but not in the nucleus.