猪ALCAM基因密码子偏好性分析、蛋白结构功能预测以及组织表达谱检测

The Codon Preference Analysis of Pig ALCAM Gene,Prediction of Protein Structure and Function,and Detection of Tissue Expression Profile

为探讨猪活化白细胞黏附分子(Activated Leukocyte Cell Adhesion Molecule,ALCAM)基因的结构与功能,本研究运用EMBOSS和Codon W软件分析猪ALCAM基因密码子使用偏好性,运用生物信息学相关软件分析ALCAM蛋白的理化性质、结构功能并预测与其存在互作关系的蛋白,运用实时荧光定量的方法检测ALCAM基因在梅山猪不同组织中的表达水平。结果表明:ALCAM基因偏好使用以A/U结尾的密码子,存在28个偏好性较强的密码子(RSCU值>1),其中CGG、AGA、GGA、GCU、ACA这5个密码子的偏好性最强,猪ALCAM基因与小鼠和酵母基因组的密码子使用差异小于大肠杆菌基因组,表明小鼠和酵母更适合作为ALCAM基因的外源表达系统;猪ALCAM基因CDS区序列全长1713 bp,编码570个氨基酸,为脂溶性、不稳定的亲水性蛋白,存在一个跨膜结构域(515~537 aa)和信号肽区域(1~27 aa);该蛋白存在5个潜在的N-糖基化位点和96个特异性蛋白激酶结合位点,二级结构以无规则卷曲和延伸链为主,可能与CD6、CD44、PTPRC和CNTN4蛋白存在互作关系。组织表达谱分析显示,ALCAM基因在梅山猪不同组织中均有表达,尤其在肺和淋巴组织中呈现高表达。本研究从密码子、蛋白质结构和功能以及组织表达谱层面初步揭示了猪ALCAM基因的结构与功能,为今后深入研究猪ALCAM基因的生物学功能提供一定的理论基础。

To explore the structure and function of porcine ALCAM gene,this study focused on the coding sequence of porcine ALCAM gene,by using EMBOSS and Codon W to analysis the codon using preferences of ALCAM gene.At the same time,using bioinformatics software to analyze the physicochemical property,basic structure and function of ALCAM protein and predicting the proteins that interact with ALCAM protein.In addition,using the method of Quantitative Real-time PCR,we detected the mRNA expression level of ALCAM gene in different tissues of Meishan pigs.The results showed that the ALCAM gene had certain preference of codon usage with the terminal base A or U,there were 28 biased codons(RSCU value>1),whose biased strongly codons were CGG,AGA,GGA,GCU and ACA.At the same time,the difference in codon usage between the porcine ALCAM gene and the mouse and Saccharomyces genome are less than the E.coli genome,it showed that Mouse and yeast were more suitable as exogenous expression systems for ALCAM genes.The whole length of the coding sequence of the porcine ALCAM gene was 1713bp,and 570 amino acids were encoded.It was a fat-soluble and unstable hydrophilic protein with a transmembrane domain(515~537aa)and a signal peptide region(1~27aa).The protein had 5 potential n-glycosylation sites and 96 specific protein kinase binding sites.The secondary structure was dominated by random coil and extended strand,which may interact with CD6,CD44,PTPRC and CNTN4 proteins.The analysis of tissue expression profile showed that ALCAM gene was expressed in different tissues of Meishan pigs,especially in lung and lymphoid tissues.This study preliminarily revealed the structure and function of porcine ALCAM gene from the aspects of codon,protein structure and function,and tissue expression profile,which provided a theoretical basis for further research on the biological function of porcine ALCAM gene.