摘要
外膜脂蛋白受体LolB是革兰氏阴性菌运输脂蛋白的关键因子。采用生物信息学方法和实时荧光定量PCR技术探究水产品腐败希瓦氏菌LolB的理化性质、结构和功能及其在不同环境中的基因表达变化。结果表明:lolB基因具有较高的序列保守性;其蛋白LolB分子质量约为23 ku,是稳定的亲水性蛋白,含有信号肽,具有21个磷酸化位点,不具有跨膜结构。LolB的二级结构以无规则卷曲和β-折叠为主,三级结构呈β桶状结构。预测到LolB与脂蛋白转运系统的LolA、LolC和LolE家族跨膜蛋白、ABC转运相关蛋白及增加细胞膜刚性、维持胞壁质稳定性的催化型裂解转糖酶等多种蛋白质存在相互作用。荧光定量PCR结果表明:lolB基因在菌体受到一定程度的饥饿、乙醇、渗透压及高温胁迫时上调表达,而随着胁迫程度增加呈先升后降的表达趋势;4℃条件下,lolB基因表达模式与对照组相似;外源信号分子C6-HSL诱导lolB高表达,并和LolB蛋白的Leu51残基通过π-烷基存在相互作用。研究结果旨在为解析LolB在腐败希瓦氏菌中的功能和水产品腐败希瓦氏菌的靶向抑制提供新思路。
Outer membrane lipoprotein receptor LolB is a key factor for gram-negative bacteria to transport lipoprotein.Bioinformatics methods and real-time fluorescence quantitative PCR were used to investigate the physicochemical properties,structure and function of LolB in Shewanella putrefaciens from aquatic products and its gene expression changes under different environment.The results showed that lolB gene had high sequence conservation property and LolB was a stable hydrophilic protein with a molecular weight of about 23 ku.LolB contained signal peptides and 21 phosphorylation sites,but had no transmembrane structure.The secondary structure of LolB was mainly random coil andβ-folding,and the tertiary structure wasβbarrel-shaped.LolB was predicted to interact with a variety of proteins,including LolA of lipoprotein transport system,LolC and LolE family transmembrane proteins,ABC transport-related proteins,catalytic cracking glycotransferase involved in increasing cell membrane rigidity and maintaining cytoplasmic stability.Fluorescence quantitative PCR results showed that the expression of lolB gene was up-regulated when the bacteria were subjected to a certain degree of stravation,ethanol,osmotic pressure and high temperature stress,which presented a trend of up-regulation and then down-regulation with the increase of stress degree.At 4℃,lolB gene expression pattern was similar to that of the control group.Exogenous signal molecules C 6-HSL induced high expression of lolB and could interact with Leu51 residue of LolB protein throughπ-alkyls.It was hoped that the results could provided a preliminary analysis of the function of LolB in S.putrefaciens and offered a new idea for the targeted inhibition of S.putrefaciens in aquatic products.
作者
周文萱
李秋莹
崔方超
檀茜倩
孙彤
励建荣
ZHOU Wenxuan;LI Qiuying;CUI Fangchao;TAN Xiqian;SUN Tong;LI Jianrong(College of Food Science and Engineering/National&Local Joint Engineering Research Center of Storage,Processing and Safety Control Technology for Fresh Agricultural and Aquatic Products,Bohai University,Jinzhou 121013,China)
出处
《食品科学技术学报》
EI
CAS
CSCD
北大核心
2023年第2期131-140,共10页
Journal of Food Science and Technology
基金
辽宁省自然科学基金资助项目(2020-MS-288)。
关键词
腐败希瓦氏菌
外膜脂蛋白定位系统
外膜脂蛋白受体LolB
实时荧光定量PCR
生物信息学分析
环境胁迫
Shewanella putrefaciens
lipoprotein outer membrane localization system
outer membrane lipoprotein receptor LolB
real-time fluorescence quantitative PCR
bioinformatic analysis
environmental stress
作者简介
第一作者:周文萱,女,硕士研究生,研究方向为水产品腐败菌致腐机制。;通信作者:李秋莹,女,讲师,博士,主要从事水产品保鲜及质量安全控制方面的研究;通信作者:励建荣,男,教授,博士,博士生导师,主要从事水产品和果蔬贮藏加工及质量安全控制方面的研究。
