HemeCu Oxidase: Calculated Electron and Proton Transfers

描述（由申请人提供）：应该可以定量地了解给定蛋白质三维结构的功能特性。这项工作将分析血红素-铜氧化酶双核中心（BNC）中02还原为水所释放的能量如何驱动矢量质子转移，从而产生跨膜电化学梯度。假设是在反应中BNC键的变化，几何和/或电荷分布被传递到蛋白质的其余部分。这就改变了质子输入和出口区域的离子化基团和氢键连通性的分布。MCCE（多构象连续静电）将用于跟踪整个蛋白质的这些变化，提供：(1)反应周期中每个阶段的净蛋白质摄取（可以与测量值进行比较）和质子结合的特定位点（难以通过实验确定）。(2)蛋白质环境对反应自由能的改变和对影响化学的残基的鉴定；(3)每一步产生的跨膜能量，以及电子转移、质子转移或偶极重排对能量贡献的分解；(4)识别构象变化，这些构象变化可以作为质子泵送的入口——这些构象变化将允许质子在反应的不同阶段进入膜的不同侧面。有关BNC显式变化的信息将通过与其他进行反应化学高水平DFT分析的人员合作获得。这些变化将用于驱动目标分子动力学。合成的结构将用MCCE进行评估，以了解BNC的反应如何修饰蛋白质。计算结果将确定牛细胞色素c氧化酶与P. dinitificans和Rb中细胞色素c氧化酶的种间差异。并与同源的大肠杆菌喹啉氧化酶结合。突变的影响将被预测。将确定稳定膜蛋白内部电荷转移并将这些电子转移偶联到跨膜质子转移的蛋白质结构基序。

DESCRIPTION (provided by applicant): It should be possible to quantitatively understand functional properties of a protein given its 3-dimensional structure. The proposed work will analyze how the energy released from reduction of 02 to water in the binuclear center (BNC) of Heme-Cu oxidases drives vectorial proton transfer, generating a transmembrane electrochemical gradient. The hypothesis is that on reaction changes in BNC bonding, geometry and/or charge distribution are transmitted to the rest of the protein. This then changes the distribution of ionized groups and hydrogen bond connectivity in proton input and exit regions. MCCE (Multi-Conformation Continuum Electrostatics) will be used to follow these changes throughout the protein, providing: (1) net protein uptake at each stage in the reaction cycle (which can be compared with measured values) and specific sites where protons are bound (which is difficult to determine experimentally). (2) modifications of reaction free energies by the protein environment and identification of residues that influence chemistry; (3) transmembrane AW generated on each step and a breakdown of how much is contributed by electron transfer, proton transfer or dipolar rearrangements; (4) identification of conformational changes that could serve as gates in proton pumping - these would permit access of protons to different sides of the membrane in different stages of the reaction. Information about explicit changes in the BNC will be obtained from collaboration with others carrying out high level DFT analysis of the reaction chemistry. These changes will be used to drive targeted molecular dynamics. The resultant structures will be evaluated with MCCE to see how the reaction at the BNC modifies the protein. The calculations will determine interspecies variation comparing the bovine cytocbrome c oxidase to that found in P. dinitificans and Rb. sphaero ides and to the homologous E. coli quinol oxidase. The effects of mutation will be predicted. Protein structural motifs that stabilize charge transfers deep inside membrane proteins and couple these electron transfers to transmembrane proton transfer will be identified.