EVOLUTION OF REDOX REACTIVITY IN CYTOCHROME B

细胞色素 B 中氧化还原反应性的演变

• 批准号: 6525848
• 负责人: RACHAEL A KIPP
• 金额: $ 2.21万
• 依托单位: PRINCETON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-01 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6525848
• 关键词: biochemical evolution; chemical kinetics; chemical substitution; computer simulation; cytochrome b; flash photolysis; heme; intermolecular interaction; metalloporphyrins; model design /development; molecular dynamics; nuclear magnetic resonance spectroscopy; oxidation reduction reaction; peptide library; physical model; protein engineering; protein structure function; site directed mutagenesis; solvents; statistics /biometry; structural biology; thermodynamics

The redox potential sets the energy yield possible in metabolism, and is also a key determinant of the rate at which the redox reaction proceeds. Understanding the factors that govern redox reactivity represents a current frontier in structure-function relationships for electron transfer proteins. The heme protein cytochrome bw2 is used as a paradigm to study the co-evolution of redox potential and redox reactivity within a systematic library of variants. Function evolves by single or pair wise mutations of residues in the vicinity of the heme. Both thermodynamic potential (EO) and redox reactivity (self exchange and cross exchange rates) are measured. All possible variants (all 20 amino acids) at each position are cloned, and a simple color screen allows the mutants that have retained both structure and affinity for the b-type heme to be determined. The extrema are used as progenitors of the next generation of variants. Comparing the variation of redox potential with that of the kinetic reactivity (within the context of Marcus theory) in these mutants will give insights into the degree of evolutionary coupling between these two properties.

氧化还原电势决定了新陈代谢中可能的能量产量，也是氧化还原反应进行速度的关键决定因素。了解控制氧化还原活性的因素是电子转移蛋白结构-功能关系的当前前沿。以血红素蛋白细胞色素bw2为范例，研究了系统变异体文库中氧化还原电位和氧化还原活性的共同进化。功能的进化是通过血红素附近残基的单个或成对突变来实现的。测量了热力学势能(EO)和氧化还原反应性(自交换和交叉交换速率)。每个位置的所有可能的变体(所有20个氨基酸)都被克隆，一个简单的彩色屏幕允许确定既保留了b型血红素的结构又保留了亲和力的突变体。极值被用作下一代变种的祖先。将这些突变体中氧化还原电位的变化与动力学反应性的变化(在马库斯理论的背景下)进行比较，将有助于深入了解这两种特性之间的进化耦合程度。