扎那纹胸鮡g型溶菌酶的cDNA克隆及组织表达分析  

Molecular characterization and tissue expression of g-type lysozyme in glytothorax zainaensis

作　　者：王运娇 吕梦 蔡莎莎 Wang Yunjiao;Lv Meng;Cai Shasha(Graduate School,Shandong First Medical University(Shandong Academy of Medical Sciences),Jinan 250000)

机构地区：[1]山东第一医科大学(山东省医学科学院)研究生院,济南250000

出　　处：《安徽医科大学学报》2023年第5期711-717,共7页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:31900328);济宁医学院教育教学研究项目(编号:Y2020018)。

摘　　要：目的鉴定扎那纹胸鮡g型溶菌酶(GzLysG)编码序列,分析其在宿主免疫系统中的作用。方法通过巢氏PCR技术克隆了GzLysG cDNA序列,通过ExPASy、SignalP 5.0、CDD、Clustal Omega等在线软件对GzLysG蛋白进行生物信息学分析,利用RT-qPCR技术分析了GzLysG mRNA在不同组织中的表达情况。结果GzLysG cDNA的开放阅读框全长558 bp,编码185个氨基酸,其蛋白序列分子量为20478.20 u,理论等电点为9.16,是一种稳定的碱性亲水性蛋白质。该蛋白无信号肽,含有g型溶菌酶典型的催化活性位点、GEWL结构域和SLT结构域。高级结构分析发现,GzLysG蛋白以α螺旋和无规则卷曲结构为主,其分子表面有一条与溶菌酶活性密切相关的狭长裂缝。系统进化分析表明溶菌酶在物种进化过程中较为保守,且GzLysG与斑马鱼来源的溶菌酶表现出较近的亲缘关系。组织表达谱分析发现,GzLysG mRNA在免疫保护相关的皮肤、肌肉和鳃组织中具有较高表达量。结论GzLysG在扎那纹胸鮡固有免疫防御中可能发挥重要作用。Objective To identify and characterize the g-type lysozyme in G.zanaensis and analyze its role in host immune system.Methods GzLysG cDNA sequence was cloned by Nest PCR technology.Bioinformatics analysis of the GzLysG protein was carried out by ExPASy,SignalP 5.0,CDD,Cluster Omega and other online software.The GzLysG gene expression pattern in G.zanaensis tissues was detected by RT-qPCR.Results The results showed that open reading frame of GzLysG cDNA was 558 bp in length,encoding 185 amino acids.The molecular weight and theoretical isoelectric point of the reduced protein was 20478.20 and 9.16,respectively.GzLysG was predicted to be a basic and hydrophilic protein,had no signal peptide and contained the typical catalytic active site,GEWL domain and SLT domain of g-type lysozyme.Advanced structural analysis revealed that GzLysG protein was mainly composed ofα-helix and random coil.There was a long and narrow crack on GzLysG molecular surface which was closely related to lysozyme activity.Lysozyme was highly conserved in evolution,with GzLysG showing a close topologic relationship with lysozyme from Danio rerio.Quantitative real-time polymerase chain reaction analysis indicated that GzLysG ubiquitously existed in all examined tissues,with higher mRNA expression levels observed in skin,muscle and gill.Conclusion All those results suggest that GzLysG plays a key role in G.zanaensis immune defensive system.

关 键 词：扎那纹胸鮡 g型溶菌酶 生物信息学分析 组织表达谱 

分 类 号：Q956[生物学—动物学]