胡萝卜软腐果胶杆菌CpxP蛋白纯化及抑菌功能鉴定

Purification and bacteriostatic identification of CpxP protein from Pectobacterium carotovorum subsp. carotovorum

摘要:

胡萝卜软腐果胶杆菌是世界十大植物病原菌之一，主要侵染十字花科的经济作物和观赏花卉。文中从胡萝卜软腐果胶杆菌的基因组中克隆1个抗菌基因cpxP (Gene ID: 29704421)，将其构建在原核表达质粒pET-15b上，并转化至大肠杆菌Escherichia coli BL21 (DE3)进行表达，经纯化后进行稳定性和抑菌实验。结果显示，IPTG的诱导终浓度为1 mmol/L，实现了蛋白的高效外源表达，纯化后电泳无杂蛋白残留，且该蛋白具有良好的热稳定性和pH稳定性。CpxP蛋白抑菌试验结果显示其对胡萝卜切片的抑菌率可达到44.89%，对马铃薯切片的抑菌率可达到59.41%。为进一步解释其抑菌机理，研究该蛋白的空间结构可为软腐病的防治和新型蛋白农药靶点研究提供新思路。

Abstract:

Pectobacterium carotovorum subsp. carotovorum is one of the world’s top ten plant pathogens, mainly infecting cruciferous economic crops and ornamental flowers. In this study, an antibacterial gene cpxP (Gene ID: 29704421) was cloned from the genome of Pectobacterium carotovorum subsp. carotovorum, and constructed on the prokaryotic expression plasmid pET-15b, and the recombinant plasmid was transformed into Escherichia coli BL21 (DE3), then stability and bacteriostatic experiments of the purified CpxP protein were performed. The final concentration of IPTG was 1 mmol/L, obtaining high-efficiency exogenous expression of the CpxP protein. There was no other protein after purification, and the destined protein exhibited good thermal stability and pH stability. The antibacterial test results showed that the inhibition rate of the CpxP protein on carrot slice was 44.89% while the inhibition rate on potato slice was 59.41%. To further explain its antibacterial mechanism, studying the spatial structure of this protein can provide new ideas for the control of soft rot and new protein pesticide targets.