Genetic and biochemical characterization of protein complexes that are essential for bacterial chromosome segregation and maintenance

蛋白质-蛋白质相互作用(分子“握手”)对所有生物的运作和功能至关重要。这些相互作用界面通常是由来自每个蛋白质伙伴的关键氨基酸残基的子集形成的。界面残基在不同的直系物之间经常不同，表明某种程度的易变性或简并性。但目前尚不清楚界面的可塑性有多大，以及界面上关键氨基酸残基的结合如何支持强大而特定的蛋白质-蛋白质相互作用。这个DTP博士项目旨在全面描述一种祖先蛋白PARB周围的序列空间，这种蛋白质对细菌染色体分离至关重要。学生将：(I)通过饱和突变和深度测序，描述对PARB-PARB相互作用至关重要的所有可能的突变和氨基酸子集上的突变组合对功能的影响。(Ii)通过体外和体内技术的结合，验证已识别的PARB变体的功能。(Ii)将突变序列空间投影到PARB的三维结构上，以充分表征序列-结构-功能关系。我们使用新月桂杆菌染色体分割蛋白B(PARB)作为该项目的理想模型系统。

Protein-protein interactions (molecular "handshakes") are crucial for the operation and function of all living organisms. These interaction interface are often formed by a subset of key amino acid residues from each protein partner. Interfacial residues often vary between orthologs, indicating some degree of mutability or degeneracy. But it is unclear how plastic the interface is and how the combination of key amino acid residues at the interface supports a robust and specific protein-protein interaction. This DTP PhD project aims to comprehensively characterise the sequence space around an ancestral protein ParB that is crucial for bacterial chromosome segregation. The student will:(i) characterise the effect on functions of all possible mutations and combination of mutations at a subset of amino acids that are crucial for ParB-ParB interaction by saturated mutagenesis and deep sequencing.(ii) validate the functionality of the identified ParB variants by a combination of in vitro and in vivo techniques.(ii) project the mutational sequence space on to the three-dimensional structure of ParB to fully characterise the sequence-structure-function relationship. We use the Caulobacter crescentus chromosome partitioning protein B (ParB) as an ideal model system for this project.