Ivancich A, Artz K, Williams J C, Allen J P, Mattioli T A
Section de Biophysique des Protéines et des Membranes, Département de Biologie Cellulaire et Moléculaire, CEA and URA 2096, Gif-sur-Yvette, France.
Biochemistry. 1998 Aug 25;37(34):11812-20. doi: 10.1021/bi9806908.
The primary donor, P, of photosynthetic bacterial reaction centers (RCs) is a dimer of excitonically interacting bacteriochlorophyll (BChl) molecules. The two constituents are named PL and PM to designate their close association with the L- and M-subunits, respectively, of the RC protein. A series of site-directed mutants of RCs from Rhodobacter sphaeroides has been constructed in order to model the effects of hydrogen bonding on the redox midpoint potential and electronic structure of P. The leucine residue at position M160 was genetically replaced with eight other amino acid residues capable of donating a hydrogen bond to the C9 keto carbonyl group of the PM BChl a molecule of P. Fourier transform (FT) (pre)resonance Raman spectroscopy with 1064 nm excitation was used to (i) determine the formation and strengths of hydrogen bonds on this latter keto carbonyl group in the reduced, neutral state (PO), and (ii) determine the degree of localization of the positive charge on one of the two constituent BChl molecules of P in its oxidized, radical cation state (P*+). A correlation was observed between the strength of the hydrogen bond and the increase in PO/P*+ redox midpoint potential. This correlation is less pronounced than that observed for another series of RC mutants where hydrogen bonds to the four pi-conjugated carbonyl groups of P were broken or formed uniquely involving histidinyl residues [Mattioli, T. A., Lin, X., Allen, J. P. and Williams, J. C. (1995) Biochemistry 34, 6142-6152], indicating that histidinyl residues are more effective in raising the PO/P*+ redox midpoint potential via hydrogen bond formation than are other hydrogen bond-forming residues. In addition, an increase in positive charge localization is correlated with the strength of the hydrogen bond and with the PO/P*+ redox midpoint potential. This latter correlation was analyzed using an asymmetric bacteriochlorophyll dimer model based on Hückel-type molecular orbitals in order to obtain estimates of certain energetic parameters of the primary donor. Based on this model, the correlation is extrapolated to the case of complete localization of the positive charge on PL and gives a predicted value for the P/P+ redox midpoint potential similar to that experimentally determined for the Rb. sphaeroides HL(M202) heterodimer. The model yields parameters for the highest occupied molecular orbital energies of the two BChl a constituents of P which are typical for the oxidation potential of isolated BChl a in vitro, suggesting that the protein, as compared to many solvents, does not impart atypical redox properties to the BChl a constituents of P.
光合细菌反应中心（RC）的主要供体P是激子相互作用的细菌叶绿素（BChl）分子的二聚体。这两个组分分别命名为PL和PM，以表明它们分别与RC蛋白的L亚基和M亚基紧密相连。为了模拟氢键对P的氧化还原中点电位和电子结构的影响，构建了一系列来自球形红杆菌RC的定点突变体。M160位的亮氨酸残基被其他八个能够向P的PM BChl a分子的C9酮羰基提供氢键的氨基酸残基通过基因方式取代。使用1064 nm激发的傅里叶变换（FT）（预）共振拉曼光谱来（i）确定还原态、中性态（PO）下该后一个酮羰基上氢键的形成和强度，以及（ii）确定P的氧化态、自由基阳离子态（P*+）下两个组成BChl分子之一上正电荷的局域化程度。观察到氢键强度与PO/P*+氧化还原中点电位的增加之间存在相关性。这种相关性不如在另一系列RC突变体中观察到的明显，在该系列突变体中，与P的四个π共轭羰基的氢键被破坏或形成，且唯一涉及组氨酸残基[马蒂奥利，T. A.，林，X.，艾伦，J. P.和威廉姆斯，J. C.（1995年）《生物化学》34卷，6142 - 6152页]，这表明组氨酸残基通过形成氢键提高PO/P*+氧化还原中点电位比其他形成氢键的残基更有效。此外，正电荷局域化的增加与氢键强度以及PO/P*+氧化还原中点电位相关。使用基于休克尔型分子轨道的不对称细菌叶绿素二聚体模型分析了后一种相关性，以便获得主要供体某些能量参数的估计值。基于该模型，将相关性外推到正电荷完全局域在PL上的情况，并给出了P/P+氧化还原中点电位的预测值，类似于对球形红杆菌HL（M202）异二聚体实验测定的值。该模型得出了P的两个BChl a组分的最高占据分子轨道能量的参数，这些参数对于体外分离的BChl a的氧化电位来说是典型的，这表明与许多溶剂相比，蛋白质并没有赋予P的BChl a组分非典型的氧化还原性质。
