小鼠CYP2J5基因组织差异表达及蛋白质结构与功能分析

Tissue differential expression and analysis of protein structure and function of mouse CYP2J5 gene

为探讨小鼠CYP2J5的结构特征及生物学功能,首先通过实时荧光定量PCR分析了小鼠CYP2J5基因组织差异表达,然后采用生物信息学工具进行了CYP2J5蛋白质结构与功能的预测和分析;为进一步探索和验证CYP2J5生物学功能及对脂代谢和性激素合成的影响,基于基因工程技术构建CYP2J5基因的超表达载体,并通过Western Blot确认构建的载体能正确表达CYP2J5基因。最终将构建的CYP2J5超表达载体导入小鼠TM3细胞,通过qPCR分析了脂代谢和性激素合成相关基因表达。结果显示,小鼠CYP2J5基因在肝脏和肾脏中高水平表达,该基因属可溶性胞质蛋白质,蛋白质二、三级结构主要以α-螺旋和无规则卷曲为主,有信号肽存在并在第13~35及75~97个氨基酸之间存在跨膜区,其主要功能是参与细胞脂代谢和脂肪酸代谢等生物调节过程。在TM3细胞中超表达CYP2J5基因后,脂代谢与性激素通路关键基因有显著变化。结果为CYP2J5基因功能的研究与开发利用提供了生物信息学和分子生物学方面的基础数据。

This study firstly analyzed the tissue differential expression of CYP2J5 gene by qRT-PCR,and then used bioinformatics tools to predict and analyze the structure and function of mouse CYP2J5.To further explore and verify the biological function and its effects on lipid metabolism and sex hormone synthesis,the overexpression vector of CYP2J5 gene was constructed based on genetic engineering technology,and the constructed vector was confirmed by Western blot which can express CYP2J5 gene correctly.Finally,the constructed CYP2J5 overexpression vector was introduced into TM3 cells,and the expression of lipid metabolism and sex hormone synthesis related genes were analyzed by qRT-PCR.The results showed that the gene was highly expressed in the liver and kidney,and it was a soluble cytoplasmic protein.The secondary and tertiary structure of the protein was mainly composed ofα-helix and random coil,the signal peptide existed and there was a transmembrane region between the 13 th to 35 th and 75 th to 97 th amino acids,whose main functions were involved in biological regulation processes such as cell lipid metabolism and fatty acid metabolism.After overexpression of the gene in TM3 cells,the key genes in the lipid metabolism and sex hormone pathways were significantly altered.The results of this study provid basic data on bioinformatics and molecular biology for the research and development gene function of CYP2J5.