摘要
【目的】瓦雷兹芽孢杆菌(Bacillus velezensis)FZB42中的未知蛋白CcdC含有一个未知结构和功能的DUF1453结构域。利用生物信息学方法预测CcdC的生物学特性,并通过构建ccdC敲除株,研究CcdC蛋白的功能,为揭示DUF1453的结构和功能提供基础。【方法】利用生物信息学在线软件预测CcdC蛋白质的氨基酸组成及理化性质、疏水性、跨膜区、亚细胞定位等,并对不同细菌中的CcdC蛋白进行系统进化树分析。构建瓦雷兹芽孢杆菌FZB42的ccdC敲除株,比较野生型FZB42与ccdC敲除株在LB液体及固体培养基上的生长情况和DSM培养基中的芽孢形成情况。【结果】CcdC蛋白由160个氨基酸组成,它是一个位于细胞膜上且无信号肽的碱性、稳定、疏水的跨膜蛋白,存在3个丝氨酸磷酸化位点,其中第7位最有可能发生磷酸化。二级结构分析显示,CcdC蛋白主要以α-螺旋为主;三级结构分析显示,现有数据库中不存在与CcdC结构相似的蛋白质。通过菌落PCR,确定ccdC敲除株构建成功。比较FZB42与ccdC敲除株在生长和芽孢形成上的差异,发现ccdC基因的缺失影响了细菌生长和芽孢形成。【结论】生物信息学分析初步确定CcdC是一个膜蛋白,磷酸化位点的预测表明其可能参与信号传递功能,相互作用网络推测CcdC蛋白可能与生长发育有关。通过构建敲除菌株,明确了ccdC基因的缺失会影响细菌的生长发育,为后续探究DUF1453结构域的生物学功能和CcdC对瓦雷兹芽孢杆菌FZB42的生长发育调控奠定了基础。
【Objective】The CcdC protein in Bacillus velezensis FZB42 contains a DUF1453 domain with an unknown structure and function.This study aims to predict the biological properties of CcdC using bioinformatics methods and to investigate its function by constructing a ccdC knockout strain,thereby laying the groundwork for elucidating the structure and function of the DUF1453 domain.【Method】Bioinformatics online tools were employed to predict the amino acid composition,physicochemical properties,hydrophobicity,transmembrane regions,and subcellular localization of the CcdC protein.A phylogenetic tree analysis of CcdC proteins from different bacterial species was also conducted.A ccdC knockout strain of B.velezensis FZB42 was constructed,and the growth of the wild-type FZB42 strain and the ccdC knockout strain was compared in both liquid and solid LB media.Additionally,spore formation of both strains was assessed in DSM medium.【Result】The CcdC protein consisted of 160 amino acids and was a basic,stable,hydrophobic transmembrane protein located in the cell membrane,lacking a signal peptide.Secondary structure analysis revealed that the CcdC protein was predominantlyα-helical.Tertiary structure analysis indicated that no structurally similar proteins to CcdC existed in current databases.Colony PCR confirmed the successful construction of the ccdC knockout strain.Comparative analysis of growth and spore formation between the wild-type FZB42 and the ccdC knockout strain demonstrated that the deletion of the ccdC gene significantly affected bacterial growth and spore formation.【Conclusion】Bioinformatics analysis tentatively identified CcdC as a membrane protein,with predicted phosphorylation sites suggesting its potential involvement in signal transduction functions.Interaction network analysis indicated that CcdC protein may be related to growth and development.The construction of the knockout strain confirmed that the deletion of the ccdC gene impair bacterial growth and development,providing a foundation for further research into the biological function of the DUF1453 domain and the regulatory role of CcdC in the growth and development of B.velezensis FZB42.
作者
李美橘
沈紫竹
官陈郧
樊奔
赵银娟
LI Meiju;SHEN Zizhu;GUAN Chenyun;FAN Ben;ZHAO Yinjuan(Co-Innovation Center for Sustainable Forestry in Southern China,Nanjing 210037,China;College of Forestry and Grassland,College of Soil and Water Conservation,Nanjing Forestry University,Nanjing 210037,China;College of Life Sciences,Nanjing Forestry University,Nanjing 210037,China)
出处
《南京林业大学学报(自然科学版)》
北大核心
2025年第6期247-254,共8页
Journal of Nanjing Forestry University:Natural Sciences Edition
基金
江苏高校优势学科建设工程资助项目(PAPD)。
