盘基网柄菌基网柄菌HtrA2基因的克隆与生物信息学分析

Cloning and Bioinformatics Analysis of Dictyostelium Discoideum HtrA2

HtrA2在细胞凋亡、细胞应激反应和神经退行性病变方面起重要作用,但其机理尚不清楚。本研究以盘基网柄菌作为模式生物,利用PCR技术克隆了盘基网柄菌HtrA2基因,经双酶切鉴定和序列测定确认后,对其进行了生物信息学分析。结果表明:所克隆盘基网柄菌HtrA2基因全长2023 bp,插入方向正确,无任何突变或碱基缺失。盘基网柄菌HtrA2蛋白分子量为72.17 kD,其分子式为C3180H5047N895O996S13,理论等电点为5.90,含257个非极性氨基酸和390个极性氨基酸。该蛋白不稳定指数为39.03,其结构相对稳定。脂溶指数和总疏水性平均值分别为94.5和-0.343。盘基网柄菌HtrA2蛋白含跨膜结构和信号肽识别序列,存在明显PDZ结构域。二级结构预测发现HtrA2蛋白含α-螺旋142个,β-转角41个,无规卷曲302个,延伸链162个,其分别占比为21.95%、6.33%、46.68%和25.04%。上述研究为后期探究HtrA2蛋白的功能与线粒体功能紊乱的相关性及其在帕金森病和其他疾病发生方面的作用机制奠定了基础。

HtrA2 play crucial roles in cell apoptosis,cell stress response and neurodegenerative disease,but its mechanisms remain unclear.Using Dictyostelium discoideum as a model,D.discoideum HtrA2 was amplified by PCR.Verified by double enzymes digestion and gene sequence,bioinformatics analysis of D.discoideum HtrA2 was then followed.The results show that D.discoideum HtrA2 gene is 2023 bp long and in the right insertion,no any mutations or base pairs loss.The molecular weight of D.discoideum HtrA2 protein is 72.17 kD and its molecular formula is C3180H5047N895O996S13.The theoretical isoelectric point of HtrA2 protein is 5.90 and the protein contains 257 nonpolar amino acids and 390 polar amino acids.The instability index of D.discoideum HtrA2 is 39.03,which means its structure is relatively stable.Its Aliphatic index and grand average of hydropathicity(GRAVY)are 94.5and-0.343 respectively.In addition,HtrA2 protein contains transmemebrane structure and signaling recognization sequence,and includes PDZ domain.The prediction of D.discoideum HtrA2 protein’s secondary structure shows that it contains 142 a-helix,41β-turns,302 random coins and 162 elongation strands,which occupies 21.95%,6.33%,46.68%and 25.04%respectively.These results shed the light on the future study to explore the relativities of the roles of HtrA2 and mitochondrial dysfunction,and its roles in PD and other human diseases.