BIOCHEMICAL-CHARACTERIZATION AND ELECTRON-TRANSFER REACTIONS OF SYM1, A RHODOBACTER-CAPSULATUS REACTION CENTER SYMMETRY MUTANT WHICH AFFECTS THE INITIAL ELECTRON-DONOR
BIOCHEMICAL-CHARACTERIZATION AND ELECTRON-TRANSFER REACTIONS OF SYM1, A RHODOBACTER-CAPSULATUS REACTION CENTER SYMMETRY MUTANT WHICH AFFECTS THE INITIAL ELECTRON-DONOR
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DOI：
10.1021/bi00157a024
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发表时间：
1992-10-27
期刊：
影响因子：
2.9
通讯作者：
WOODBURY, NW
中科院分区：
文献类型：
--
作者：
TAGUCHI, AKW;STOCKER, JW;WOODBURY, NW
将编码M187-M203氨基酸的荚膜红杆菌光合作用反应中心M亚基基因(pufM结构序列的M562-M612核苷酸)的51bp片段替换为L亚基基因的同源区。这导致了反应中心初始电子供体P的大部分氨基酸环境的对称化。这是一系列大规模对称突变中的第一个，被称为sym1。Sym1突变体能够以光合作用方式生长，表明反应中心功能基本完好。分离的反应中心在P的Q(Y)带有大约10 nm的蓝移。通过分析苯醌还原反应中心的长寿命荧光，确定了P*和P+BPH(A)-之间的标准自由能变化，从野生型的约-120 meV下降到sym1突变体的约-75 meV。观察到从P*到P+Q(A)-的电子转移量子产额为65-70%，大部分产额损失发生在P*和P+BPH(A)-之间。该突变体的受激辐射衰减率约为野生型的3倍。对SYM1反应中心形成的电荷分离中间体的时间分辨光谱分析表明，主要产物为P+BPH(A)-。对观察到的速率和电子转移产率的模型相关分析给出了sym1反应中心的以下微观速率常数(相同条件下的野生型值在括号中)：[图形]对sym1突变体、其他基团在P附近产生的突变体以及P附近氨基酸的种间变异的分析表明，初始电子供体环境中的蛋白质不对称性对于优化电子转移的速率和产率是重要的，但对整个反应中心的功能并不是严格要求的。
A 51 bp section of the Rhodobacter capsulatus photosynthetic reaction center M subunit gene (nucleotides M562-M612 of the pufM structural sequence) encoding amino acids M187-M203 was replaced by the homologous region of the L subunit gene. This resulted in the symmetrization of much of the amino acid environment of the reaction center initial electron donor, P. This is the first in a series of large-scale symmetry mutations and is referred to as sym1. The sym1 mutant was able to grow photosynthetically, indicating that reaction center function was largely intact. Isolated reaction centers showed an approximately 10-nm blue shift in the Q(Y) band of P. The standard free energy change between P* and P+Bph(A)- determined from analysis of the long-lived fluorescence from quinone-reduced reaction centers decreased from about -120 meV in the wild-type to about -75 meV in the sym1 mutant. A 65-70% quantum yield of electron transfer from P* to P+Q(A)- was observed, most of the yield loss occurring between P* and P+Bph(A)-. The decay of the stimulated emission from P* was about 3-fold slower in this mutant than in the wild-type. Time-resolved spectral analysis of the charge-separated intermediates formed in sym1 reaction centers indicated that the major product was P+Bph(A)-. A model-dependent analysis of the observed rates and electron-transfer yields gave the following microscopic rate constants for sym1 reaction centers (wild-type values under the same conditions are given in parentheses):[GRAPHICS]Analysis of the sym1 mutant, mutants near P made by other groups, and interspecies variation of amino acids in the vicinity of P suggests that the protein asymmetry in the environment of the initial electron donor is important for optimizing the rate and yield of electron transfer, but is not strictly required for overall reaction center function.