Large-scale mapping of the protein function landscape

蛋白质功能图谱的大规模绘图

• 批准号: 8686612
• 负责人: Philip Anthony Romero
• 金额: $ 5.33万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8686612
• 关键词: Abate; Affect; Amino Acid Sequence; Biological Assay; DNA; DNA Sequence; Data; Disease; Engineering; Enzymes; Evolution; Gene Mutation; Genetic Epistasis; Genomics; Goals; Hereditary Disease; High-Throughput DNA Sequencing; Hour; Human; Human Genetics; Hyperargininemia; In Vitro; Individual; Knowledge; Laboratories; Lead; Malignant Neoplasms; Maps; Medicine; Methods; Microfluidic Microchips; Microfluidics; Modeling; Molecular; Mutation; Nature; Pathway interactions; Peptide Sequence Determination; Property; Protein Engineering; Proteins; Research; Role; Schools; Statistical Models; Technology; Testing; Therapeutic; Variant; arginase; base; biophysical properties; chemotherapeutic agent; design; high throughput screening; insight; interest; predictive modeling; protein function; public health relevance; sequence learning; urea cycle

DESCRIPTION (provided by applicant): Knowledge of how mutations affect protein function is important for understanding how natural proteins evolved, engineering new proteins with useful properties, and predicting disease-associated mutations. Epistasis is the phenomenon where the effect of one mutation depends on the presence of another mutation. These epistatic interactions stymie our ability to predict the phenotypic consequences of mutations, and can restrict the mutational pathways that are available to evolution. Laboratory evolution studies have highlighted the multifaceted nature of protein function and how competing biophysical properties, such as enzymatic activity and protein stability, can generate epistatic interactions between residues. I hypothesize that most epistasis arises as a result of these pleiotropic mechanisms, rather than direct physical interactions between residues. The human urea cycle enzyme Arginase I (hArgI) will be used to study how sequence changes affect expression, stability, and enzymatic activity. The experimental approach will leverage recent advances in parallel DNA sequencing and ultra-high- throughput screening to map the protein function landscape on an unprecedented scale. These data will be used to explore how combinations of biophysical properties generate mutational epistasis and define the space of functional protein sequences.

描述（由申请人提供）：有关突变如何影响蛋白质功能的知识对于理解天然蛋白的发展方式很重要，具有有用特性的新蛋白质以及预测与疾病相关的突变很重要。上科斯是一种现象，其中一个突变的作用取决于另一个突变的存在。这些上任的相互作用阻碍了我们预测突变的表型后果的能力，并可以限制可用于进化的突变途径。实验室进化研究强调了蛋白质功能的多方面性质，以及诸如酶活性和蛋白质稳定性之类的竞争性生物物理特性如何在残基之间产生上皮相互作用。我假设由于这些多效机制而不是直接残基之间的物理相互作用，大多数上毒是由于这些多效机制而产生的。人尿素周期精氨酸酶I（HARGI）将用于研究序列变化如何影响表达，稳定性和酶活性。实验方法将利用平行DNA测序和超高吞吐量筛选的最新进展，以绘制前所未有的量表。这些数据将用于探索生物物理特性的组合如何产生突变的上毒并定义功能蛋白序列的空间。