A genomic and structural study of FtsZ constriction in cell division

细胞分裂中 FtsZ 收缩的基因组和结构研究

• 批准号: 8269666
• 负责人: Kiani Anela Jeniah Arkus Gardner
• 金额: $ 2.94万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2013-05-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8269666
• 关键词: Antibiotics; Bacteria; Bacterial Proteins; Base Pairing; Binding Sites; Bioinformatics; C-terminal; Caulobacter; Cell division; Cell membrane; Cells; Chromosome Mapping; Cleaved cell; Complement; Coupled; DNA Resequencing; Data; Dideoxy Chain Termination DNA Sequencing; Engineering; Escherichia coli; Fluorescence Microscopy; Frequencies; Generations; Genes; Genome; Genomics; Goals; Homologous Gene; In Vitro; Lateral; Length; Link; Liposomes; Maps; Membrane; Modeling; Mutation; Parents; Pathway interactions; Phenotype; Point Mutation; Prokaryotic Cells; Proteins; Rhizobium; Role; Site; Structure; Suppressor Mutations; System; TEV protease; Tail; Technology; Temperature; Testing; Thick; Tubulin; Variant; X-Ray Crystallography; adducin; constriction; flexibility; in vivo; insight; interest; large scale production; loss of function; loss of function mutation; mutant; novel; plasma protein Z; programs; public health relevance; reconstitution; structural genomics

DESCRIPTION (provided by applicant): FtsZ (filamentous temperature sensitive Z) is an essential protein for bacterial cell division. It assembles the Z-ring that pinches the cell in two. I am interested in two different aspects of FtsZ function. 1. FtsZ is the only protein necessary for constriction force generation in liposomes in vitro. FtsZ from foreign bacteria and some mutant FtsZ were also able to cause cell division in E. coli. However, some forms of FtsZ tested required the acquisition of unknown genomic mutations to suppress the defective FtsZ phenotype and allow division. The mutations in these suppressor strains are currently unknown, but are likely loss-of- function mutations in other pathways involved in cell division. This project aims to utilize high-throughput sequencing technology to resequence the entire genome of each of the 13 suppressor strains to map the suppressor mutation and identify further genes of interest in bacterial cell division. 2. The FtsZ protein has a C-terminal (Ct) tail region that lacks a lacks a well-defined structure as visualized through X-ray crystallography, indicating that it serves primarily as a spacer to link the FtsZ globular domain to the FtsA binding site and the membrane. However, some FtsZ mutants with small 20aa insertions in the Ct tail are incapable of functioning for cell division. This project aims to examine the role of the Ct tail function in vivo and in vitro. The Ct tail will be expressed and purified apart from the FtsZ globular domain for NMR structure determination. Varied forms of FtsZ will be expressed using alternative Ct tail sequences and length, and the ability of the protein to produce a constriction force in liposomes in vitro, and to function for cell division in vivo will be assessed. Public Health Relevance: FtsZ is a required protein for cell division in most bacteria. A detailed understanding of FtsZ structure and function would increase the possibility and efficacy of targeting the protein pharmacologically to inhibit bacterial division, thus creating potential new antibiotics.

描述（由申请人提供）：FtsZ（丝状温度敏感Z）是细菌细胞分裂的必需蛋白。它组装Z形环将细胞夹成两半。我对FtsZ函数的两个不同方面感兴趣。1. FtsZ是脂质体在体外产生收缩力所必需的唯一蛋白质。外源FtsZ和一些突变体FtsZ也能引起大肠杆菌细胞分裂。杆菌然而，测试的某些形式的FtsZ需要获得未知的基因组突变以抑制缺陷的FtsZ表型并允许分裂。这些抑制菌株中的突变目前尚不清楚，但可能是参与细胞分裂的其他途径中的功能丧失突变。该项目旨在利用高通量测序技术对13种抑制菌株中的每一种的全基因组进行重新测序，以定位抑制突变并鉴定细菌细胞分裂中感兴趣的进一步基因。2. FtsZ蛋白具有C-末端（Ct）尾区，其缺乏通过X射线晶体学观察到的明确定义的结构，表明其主要用作将FtsZ球状结构域连接到FtsA结合位点和膜的间隔区。然而，一些在Ct尾部具有小的20个氨基酸插入的FtsZ突变体不能用于细胞分裂。本项目旨在研究Ct尾功能在体内和体外的作用。表达Ct尾，并与FtsZ球状结构域分离纯化，用于NMR结构测定。将使用替代的Ct尾序列和长度表达不同形式的FtsZ，并将评估蛋白质在体外脂质体中产生收缩力和在体内发挥细胞分裂功能的能力。 公共卫生相关性：FtsZ是大多数细菌细胞分裂所需的蛋白质。对FtsZ结构和功能的详细了解将增加靶向蛋白质抑制细菌分裂的可能性和有效性，从而创造潜在的新抗生素。