徐凯^1,2, 刘鑫玟^1,2, 覃昊^1,2, 张珍珍^1,2, 阎斌伦^1,2, 高焕^1,2, 胡广伟^1,2
1.江苏海洋大学 江苏省海洋生物资源与环境重点实验室/江苏省海洋生物技术重点实验室, 江苏 连云港 222005;2.江苏海洋大学 江苏省海洋生物产业技术协同创新中心, 江苏 连云港 222005

摘要:
为研究脊尾白虾（Exopalaemon carinicauda）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。
关键词:脊尾白虾抗菌肽Crustin基因克隆原核表达
DOI：10.11759/hykx20210721002
分类号:S945.1
基金项目:国家自然科学基金（31900370）；江苏省自然科学基金（BK20191007）；连云港市“521高层次人才培养工程”（2021-1021）；江苏省海洋生物技术重点实验室开放课题（HS2019001）；江苏省研究生科研创新计划（KYCX20_2890，KYCX2021-036）

Cloning, expression, and functional analyses of Crustin in Exopalaemon carinicauda
XU Kai^1,2, LIU Xin-wen^1,2, QIN Hao^1,2, ZHANG Zhen-zhen^1,2, YAN Bin-lun^1,2, GAO Huan^1,2, HU Guang-wei^1,2
1.Jiangsu Key Laboratory of Marine Bioresources and Environment/Jiangsu Key Laboratory of Marine Biotechnology School of Marine Science and Fisheries, Jiangsu Ocean University, Lianyungang 222005, China;2.Co-Innovation Center of Jiangsu Marine Bio-industry Technology, Jiangsu Ocean University, Lianyungang 222005, China
Abstract:
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.
Key words:Exopalaemon carinicaudaantimicrobial peptidesCrustingene cloningprokaryotic expression





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