枯草芽胞杆菌异戊二烯酶ComQ的生物信息学分析及对生物膜形态的影响

Bioinformatics analysis of Bacillus subtilis isoprene enzyme ComQ and its effect on biofilm morphology

摘要:

【目的】枯草芽胞杆菌ComQ是一种类异戊二烯生物合成酶。利用生物信息学预测分析了ComQ的生物学特性，对comQ基因进行过表达和敲除，构建突变菌，孔板发酵培养验证生物膜形态变化。【方法】运用NCBI （National Center for Biotechnology Information）网站里的Protein数据库获取ComQ蛋白氨基酸序列，通过在线生物信息学软件预测分析其生物学特征，包括其理化性质、信号肽、结构域、空间结构等。构建枯草芽胞杆菌BS168的comQ过表达及敲除菌株，利用孔板发酵培养验证生物膜生长性状。【结果】ComQ蛋白由299个氨基酸组成，理论相对分子质量约34 204.08 Da，无信号肽，无跨膜区，为稳定胞内蛋白。通过菌落PCR，验证枯草芽胞杆菌BS168突变菌构建成功。利用孔板发酵培养枯草芽胞杆菌BS168及突变菌株，验证ComQ蛋白对枯草芽胞杆菌生物膜的形态形成存在影响。两种突变菌株生物膜形态均与原始菌株存在差异。【结论】通过对ComQ蛋白的基本性质、关键氨基酸位点及蛋白结构预测分析，构建突变菌株，验证生物膜形态变化，为后续探究ComQ对枯草芽胞杆菌的生长代谢变化奠定了基础。

Abstract:

[Objective] Bacillus subtilis ComQ is a kind of isoprene biosynthesis enzyme. We used bioinformatics to predict and analyze the biological characteristics of ComQ. In addition, we also constructed comQ overexpression and knockout mutants to verify their effect on biofilm morphology. [Methods] Firstly, we obtained the amino acid sequence of ComQ protein from the Protein database in the NCBI website. Secondly, we predicted and analyzed the biological characteristics including physical and chemical properties, signal peptide, domain and spatial structure of ComQ by online bioinformatics software. Thirdly, we constructed comQ overexpression and knockout strain of Bacillus subtilis, and observe their growth characteristics of biofilm by plate fermentation. [Results] ComQ protein was composed of 299 amino acids with a theoretical molecular weight of about 34 204.08 Da. It was a stable intracellular protein without signal peptide and transmembrane region. Colony PCR verified that Bacillus subtilis BS168 mutants were constructed successfully. The biofilm morphology of the two mutants was different from that of the original strain BS168. [Conclusion] Through the prediction and analysis of the basic properties, key amino acid sites and protein structure of ComQ protein, the mutant strain was constructed to verify the morphological changes of biofilm, which laid a foundation for the subsequent study of ComQ on the growth and metabolism of Bacillus subtilis.