意大利蜜蜂化学感受蛋白AmelCSP2序列分析及表达特性研究

Sequence analysis and expression characteristics of chemosen⁃ sory protein AmelCSP2 in Apis mellifera

  • 摘要: 【目的】探究意大利蜜蜂(Apis mellifera)化学感受蛋白CSP2(AmelCSP2)序列特征及AmelCSP2基因组织表达特性,为阐明AmelCSP2蛋白的生物学功能提供理论参考。【方法】利用ProtParam、ProtScal、SignalP-5.0、TMHMM-2.0、PSORTb 3.0.3、DictyOGlyc-1.1、NetNGlyc-1.0、NetPhos 3.1、SWISS-MODEL、CD-search、SMART和STRING等生物信息学软件预测AmelCSP2蛋白理化性质、亲/疏水性、信号肽、跨膜结构域、亚细胞定位、O-糖基化位点、N-糖基化位点、磷酸化位点、高级结构、模体、结构域和蛋白互作网络。采用实时荧光定量PCR检测AmelCSP2基因在意大利蜜蜂触角、头(不含触角)、胸、腹、足、翅中的相对表达量。【结果】AmelCSP2蛋白编码117个氨基酸残基,理论分子量约13.06 kD,理论等电点(pI)为9.27,分子脂肪系数为92.56,不稳定系数为59.68,平均亲水性指数为-0.254,为两性不稳定蛋白。AmelCSP2蛋白在第1~21位氨基酸残基处有信号肽,不含跨膜结构域,主要定位于细胞周质、细胞质和细胞外膜。AmelCSP2蛋白不存在糖基化位点,但存在20个磷酸化位点。AmelCSP2蛋白二级结构中无规则卷曲占42.74%,α-螺旋占38.46%,延伸链占18.80%,蛋白三级结构较稳定。AmelCSP2蛋白属于OS-D超家族,在第32~116位氨基酸残基处存在唯一蛋白结构域OS-D。AmelCSP2蛋白可能与PYX2、ASP3、OBP3c、NT-7及OBP14蛋白存在相互作用。AmelCSP2基因在意大利蜜蜂的触角、头(不含触角)、胸、腹、足、翅中均有表达,触角中AmelCSP2基因相对表达量显著高于其他组织(P<0.05)。【结论】AmelCSP2蛋白具有典型的OS-D超家族结构域,并含有多个磷酸化位点,可能通过磷酸化修饰调控其功能。AmelCSP2蛋白可能与PYX2、ASP3、OBP3c、NT-7及OBP14蛋白相互作用,参与化学感受信号传导。AmelCSP2基因在意大利蜜蜂触角中高表达,其可能在嗅觉感受系统中承担气味分子运输功能。

     

    Abstract: 【Objective】To explore the sequence characteristics of the chemosensory protein CSP2 of Apis melliferaAmelCSP2)and the tissue expression characteristics of the AmelCSP2 gene,which could provide theoretical reference for clarifying the biological function of AmelCSP2 protein.【Method】The physicochemical properties,hydrophilicity/hy‐ drophobicity,signal peptide,transmembrane domain,subcellular localization,O-glycosylation sites,N-glycosylation sites,phosphorylation sites,advanced structure,motif,domain and protein interaction network of AmelCSP2 protein were predicted by using bioinformatics softwares such as ProtParam,ProtScal,SignalP-5.0,TMHMM-2.0,PSORTb 3.0.3,DictyOGlyc-1.1,NetNGlyc-1.0,NetPhos 3.1,SWISS-MODEL,CD-search,SMART and STRING. The relative expression levels of AmelCSP2 gene in the antennae,head(excluding antennae),thorax,abdomen,leg and wing of A. mellifera were detected by real-time fluorescence quantitative PCR.【Result】The AmelCSP2 protein encoded 117 amino acid residues,with a theoretical molecular weight of approximately 13.06 kD,a theoretical isoelectric point(pI)of 9.27, a molecular fat coefficient of 92.56,an instability coefficient of 59.68,and an average hydrophilicity index of -0.254, being an amphoteric and unstable protein. The AmelCSP2 protein had a signal peptide at amino acid residues positions 1-21, did not contain transmembrane domain,and was mainly located in the periplasm,cytoplasm and extracellular membrane. The AmelCSP2 protein had no glycosylation sites but had 20 phosphorylation sites. In the secondary structure of AmelCSP2 protein,random coils accounted for 42.74%,α-helices accounted for 38.46%,and extended chains accounted for 18.80%,and the tertiary structure of the protein was relatively stable. The AmelCSP2 protein belonged to the OS-D superfamily and had the only protein domain OS-D at amino acid residues positions 32-116. The AmelCSP2 protein might interact with PYX2,ASP3,OBP3c,NT-7 and OBP14 proteins. The AmelCSP2 gene was expressed in the antennae,head (excluding antennae),thorax,abdomen,leg and wing of A. mellifera,and the relative expression level of the AmelCSP2 gene in the antennae was significantly higher than that in other tissues(P<0.05).【Conclusion】AmelCSP2 protein has a typical OS-D superfamily domain and contains multiple phosphorylation sites,which may regulate its function through phosphorylation modification. AmelCSP2 protein may interact with PYX2,ASP3,OBP3c,NT-7 and OBP14 proteins and participate in chemical sensory signal transduction. AmelCSP2 gene is highly expressed in the antennae of A. mellifera,and may play the role of odor molecule transport in the olfactory perception system.

     

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