Molecular Mechanisms of Human DNA Polymerase B Catalysis, Fidelity and Selective

人类 DNA 聚合酶 B 催化、保真度和选择性的分子机制

• 批准号: 8591712
• 负责人: MYRON GOODMAN
• 金额: $ 58.05万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-03 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8591712
• 关键词: Active Sites; Catalysis; Cell-Free System; Chemicals; DNA-Directed DNA Polymerase; Electronics; Electrostatics; Family; Grant; Human; Instruction; Kinetics; Left; Malignant Neoplasms; Measurement; Molecular; Oxygen; Polymerase; Property; Research; analog; base; bisphosphonate; cancer cell; human DNA; innovation; member; mutant; repair enzyme; research study; theories; tool

DNA polymerase mechanisms of base selection and catalysis are explored using a tool-kit of dNTP analogs that have bisphosphonates in place of the B,Y-bridge oxygen. These analogs are Pol B substrates that have leaving groups with widely dispirate electronic properties, pKa4 values ranging from 7.8 to 12.3, enabling us to use presteady state kinetic measurements to determine the selection of right and wrong deoxynucleotides occurring at the chemical transition state. Especially important members of the toolkit include all four individually synthesized (R)- and (S)-B,Y-CHF and B,Y-CHCI diastereomers. Our recent observation of a pronounced stereoselection for (R)-CHF in Pol B involving an electrostatic interaction of F with Arg183, unique to family X pols such as Pol B, serves as the impetus for a

使用dNTP类似物的工具包探索了碱基选择和催化的DNA聚合酶机制，这些dNTP类似物以双膦酸盐取代B，Y桥氧。这些类似物是POL B底物，留下具有广泛衰减电子性质的基团，pKa4值从7.8到12.3，使我们能够使用稳态前动力学测量来确定发生在化学过渡态的正确和错误脱氧核苷酸的选择。该工具包特别重要的成员包括所有四个单独合成的(R)和(S)-B，Y-CHF和B，Y-CHCI非对映异构体。我们最近观察到在POL B中存在明显的(R)-CHF的立体选择，涉及F与Arg183的静电相互作用，这是像POL B这样的X家族POL所特有的，这是导致