Kinetics and free energy gaps of electron-transfer reactions in Rhodobacter sphaeroides reaction centers.
Kinetics and free energy gaps of electron-transfer reactions in Rhodobacter sphaeroides reaction centers.
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球形红细菌反应中心电子转移反应的动力学和自由能隙。
DOI：
10.1021/bi00097a008
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发表时间：
1993
期刊：
影响因子：
2.9
通讯作者：
Schenck,CC
中科院分区：
文献类型：
--
作者：
Nagarajan,V;Parson,WW;Davis,D;Schenck,CC
19938年9月3日收到的修订稿摘要：在用苯丙氨酸、异亮氨酸或色氨酸取代酪氨酸(M)的突变菌株中，检测了球形红杆菌光合作用反应中心(RC)中光驱动的电子转移反应的速率。在0激发下，吸收光谱在700-975 nm之间变化。利用奇异值分解对605 nm的6-ps脉冲进行了全局分析。在室温下测得的光谱用一个模型解释：在野生型RCS和Phe、Lie和Trp突变体中，被激发的细菌叶绿素二聚体(P*)将一个电子转移到细菌素(Hl)上，时间常数分别为3.5±0.3、10.5±1.0、16±2和41±4ps，然后电子从hl“转移到苯二酚(Qa)，时间常数在野生型RCS中为0.16 ns，在Phe突变体中为0.24 ns，在He和Trp突变体中为0.20 ns。在野生型RCS中，第一步随着温度的下降而加快，在Phe突变体中几乎保持不变，在He和Trp突变体中减慢。在80K时，在850-975 nm区域的信号包括P~*的受激发射或吸收光谱的明显移动，在He突变体中的时间常数为5ps，而在Trp突变体中的时间常数为13ps。在He和Trp突变体中，大部分电子转移到H1的时间常数分别为55ps和155ps，可能是由P*的松弛形式发生的。然而，不能排除电子从初始态转移的可能性。在野生型RCS或Phe突变体中，P*的松弛不被分解。用电化学方法测得P/P+氧化还原电对的中点电位(Em)，野生型和Phe、He和Trp突变体RCS的中点电位分别为500±5、530±6、533±3和552±10 mV。这些值得到了化学滴定的证实。由P*形成P+Hl“自由基对的自由能变化(AG)也是通过测量阻止电子从Hl”到Qa的转移后在纳秒时间尺度上的荧光幅度来确定的。P+Hl“的自由能增加的量与He突变体中P的Em的增加相当，在Phe和Trp突变体中比这多16 meV。非绝热电子转移理论被用来将P+Hl“的形成速率常数与AG联系起来。如果假设突变只改变了整个AG，则不能充分解释突变体中反应随温度的变化，但可以通过假设它们还增加了作为动力学和虚拟中间体的附加态(P+b1”)的自由能来解释。P+Bl“自由能的必要增加量大于P+Hl”自由能的实测值，四种菌株的P+Qa“PQA回流反应均随温度降低而加快。在室温下，野生型RCS和Phe、He、Trp突变体RCS的反应时间常数分别为0.105±0.01、0.100±0.005、0.13±0.01和0.045±0.005 S。色氨酸突变体的大幅加速不能简单地用AG值的变化来解释。在紫色光合成细菌的反应中心复合体(RCS)1中，细菌-叶绿素二聚体(P)的激发导致沿两条对称位置的色素链之一的一系列电子转移反应。被激发的二聚体(P*)将一个电子转移到细菌素(Hl)上，形成一对P+hl的“自由基对”。从hl，一个电子转移到一个苯二酚(Qa)，然后再转移到第二个苯二酚(Qb)--这是…长期以来的谜团之一
Revised Manuscript Received September 3, 19938 abstract: The rates of the light-driven, electron-transfer reactions in the photosynthetic reaction center (RC) of Rhodobacter sphaeroides are examined in mutant strains in which tyrosine (M) 210 is replaced by phenylalanine, isoleucine, or tryptophan. The spectra of the absorbance changes between 700 and 975 nm, following excitation by0. 6-ps pulses at 605 nm, are analyzed globally by singular value decomposition. The spectra measured at room temperature are interpreted in terms of a model in which the excited bacteriochlorophyll dimer (P*) transfers an electron to a bacteriopheophytin (Hl) with time constants of 3.5±0.3, 10.5±1.0, 16±2, and 41±4 ps in wild-type RCs and the Phe, lie, and Trp mutants, respectively, and an electron then moves from Hl" to a quinone (Qa) with a time constant of 0.16 ns in wild-type RCs, 0.24 ns in thePhe mutant, and 0.20 ns in the He and Trp mutants. The first step speeds up with decreasing temperature in wild-type RCs, remains virtually unchanged in the Phe mutant, and slows down in the He and Trp mutants. At 80 K, thesignals in the 850-975-nm region include an apparent shift of the stimulated emission or absorption spectrum of P*, with a time constant of 5 ps in the He mutant and 13 ps in the Trp mutant. Most of the electron transfer to Hl occurs with time constants of 55 and 155 ps in the He and Trp mutants, respectively, and probably occurs from therelaxed form of P*. Electron transfer from the initial state cannot be ruled out, however. Relaxations of P* are not resolved inwild-type RCs or the Phe mutant. The midpoint potential (Em) of the P/P+ redox couple is measured by an electrochemical technique; the Em values are 500±5, 530±6, 533±3, and 552±10 mV for the wild-type and the Phe, He, and Trp mutant RCs, respectively. These values are corroborated by chemical titrations. The free energy change (AG) associated with formation of the P+ Hl" radical pair from P* also is determined by measuring the amplitude of fluorescence on the nanosecond time scale after blocking electron transfer from Hl" to Qa. The free energy of P+ Hl" is elevatedby an amount comparable to that calculated from theincrease in the Em of P in the He mutant andby about 16 meV more than this in the Phe and Trp mutants. Nonadiabatic electron-transfer theory is used to relate the rate constant of the formation of P+ Hl" to AG. The altered temperature dependence of the reaction in the mutants cannot be explained adequately on the assumption that the mutations only alter the overallAG, but it can be accounted for by assuming that they also increase the free energy of an additional state (P+ Bl") that serves as both a kinetic and a virtual intermediate. The requisite increases in the free energy of P+ Bl “are greater than the measured changes in the free energy of P+ Hl". The P+ Qa" PQa back-reaction speeds up with decreasing temperature in all four strains. At room temperature, this reaction has time constants of 0.105±0.01, 0.100±0.005, 0.13±0.01, and 0.045±0.005 s in the wild-type and the Phe, He, and Trp mutant RCs, respectively. The large acceleration in the Trp mutant cannot be explained simply in terms of the change in AGfor this reaction.In reaction center complexes (RCs) 1 of purple photosyn-thetic bacteria, excitation of a bacteriochlorophyll dimer (P) results in a series of electron-transfer reactions along one of two symmetrically positioned pigment chains. The excited dimer (P*) transfers an electron to a bacteriopheophytin (Hl), forming a P+ Hl" radical pair. From HL", an electron moves to a quinone (Qa) and then to a second quinone (Qb)-One of the longstanding puzzles …