脊尾白虾抗菌肽Crustin基因的克隆、表达与功能研究

Cloning, expression, and functional analyses of Crustin in Exopalaemon carinicauda

为研究脊尾白虾(Exopalaemoncarinicauda)抗菌肽Crustin的结构特征和功能,本研究从脊尾白虾中克隆得到一个Crustin的新变体,命名为EcCrustin1。研究结果表明,EcCrustin1基因cDNA序列长度为740 bp,开放阅读框(ORF)长度为477 bp,编码158个氨基酸,EcCrustin1蛋白具有Ⅱ型Crustin的典型特征,包括N端的信号肽,C端WAP结构域,以及二者之间的甘氨酸富集区(GRR)和半胱氨酸富集区(CRR)。EcCrustin1基因主要在脊尾白虾血淋巴中表达,副溶血弧菌刺激后,EcCrustin1在血淋巴中的表达显著上调(P<0.05),说明EcCrustin1在脊尾白虾先天免疫应答中发挥重要作用。此外,通过原核表达获得了该抗菌肽的重组蛋白,为进一步研究其抑菌功能和机理提供了基础。

A new Crustin was identified from Exopalaemon carinicauda(EcCrustin1)to reveal the structure and function of the antibacterial peptide Crustin in E.carinicauda.The results show that the length of EcCrustin1 cDNA was 740 bp,and the open reading fragment was 477 bp,which coded a 158 amino-acid polypeptide.EcCrustin1 contained a signal peptide at the N-terminus and a WAP domain at the C-terminus.There were GRR and CRR domains between the signal peptide and the WAP,showing the typical characteristics of a type II Crustin.EcCrustin1 was highly expressed in the hemolymph and was significantly upregulated(P<0.05)after a challenge with Vibrio parahaemolyticus,indicating that EcCrustin1 plays an important role in the E.carinicauda innate immune response.The recombinant protein was obtained by prokaryotic expression,which provides a reference for further study of its antibacterial function and mechanism.