噬菌体TM4中LysinB蛋白的特征及抗菌特性研究  

Characterization and antibacterial properties of LysinB protein in bacteriophage TM4

作　　者：潘稚芬 周紫薇 徐海萍 王蔚 PAN Zhifen;ZHOU Ziwei;XU Haiping;WANG Wei(Department of Tuberculosis,the First Hospital of Jiaxing,Affiliated Hospital of Jiaxing University,Jiaxing 314001,Zhejiang,China;School of Life Sciences,Fudan University,Shanghai 200438,China)

机构地区：[1]嘉兴大学附属医院嘉兴市第一医院结核科,浙江嘉兴314001 [2]复旦大学生命科学学院,上海200438

出　　处：《中国现代医生》2024年第21期6-10,25,共6页China Modern Doctor

基　　金：浙江省嘉兴市科技计划项目(2022AD30065)。

摘　　要：目的对分枝杆菌噬菌体TM4LysinB蛋白序列进行生物学分析,原核表达LysinB蛋白并评估其体外杀菌特性。方法合成噬菌体TM4 LysinB基因,构建原核表达重组质粒,诱导蛋白可溶表达、纯化,通过在线软件程序分析噬菌体TM4 LysinB蛋白的生物学特征;在7H9培养基中培养耻垢分枝杆菌,检测其杀菌效果及稳定性。结果LysinB蛋白理论等电点为6.66,不稳定指数为28.71,为稳定蛋白质,且为两亲性蛋白。二级结构中,无规则卷曲、α-螺旋、β-折叠和转角分别占42.00%、40.25%、12.00%和5.75%;三级结构中,无规则卷曲含量高,由11~73位AA组成的肽段构成肽聚糖结合结构域。预测出B细胞抗原表位区段和T细胞抗原表位的强结合肽段。LysinB蛋白具有显著的体外杀耻垢分枝杆菌的能力,pH8.0的PBST蛋白缓冲液有利于在实验周期内维持LysinB蛋白的杀菌效果。结论进一步了解分枝杆菌噬菌体TM4裂解酶LysinB蛋白,为研制可稳定贮存的噬菌体酶制剂及其临床应用提供参考。Objective To analyze the sequence of LysinB protein in bacteriophage TM4,express LysinB protein in E.coli and evaluate its bactericidal activity in vitro.Methods Bacteriophage TM4 LysinB gene was synthesized and the recombinant prokaryotic expression plasmid was constructed.The soluble protein was induced and purified.The biological characteristics of bacteriophage TM4 LysinB protein were analyzed by online software program.Mycobacterium smegmatis was cultured in 7H9 medium,and its bactericidal effect and stability were detected.Results The theoretical isoelectric point was 6.66,the instability index was 28.71,which was a stable and amphiphilic protein.In the secondary structure,The random coil,α-helix,β-fold and turn Angle accounted for 42.00%,40.25%,12.00%and 5.75%;In the tertiary structure,the content of random coil was high.The peptide composed of AA at positions 11 to 73 constituted the peptidoglycan binding domain.The regions of B cell epitopes and the strong binding peptides of T cell epitopes were predicted.LysinB protein has a significant ability to kill Mycobacterium smegmatis in vitro.PBST protein buffer with pH8.0 was beneficial to maintain the bactericidal effect of LysinB proteins during the experimental cycle.Conclusion This study expands the comprehensive understanding of the LysinB protein of mycobacterial phage TM4 lyase,and also provides reference for the development of stable storage of phage enzyme preparation and its clinical application.

关 键 词：噬菌体TM4 LysinB蛋白 耻垢分枝杆菌 生物信息学 

分 类 号：R378.91[医药卫生—病原生物学]