Controlling the Pathway of Electron Transfer in Bacterial Reaction Centers

光合反应中心内的光引发电子转移是光合生物转换太阳能的主要事件。紫色非硫细菌光合反应中心的核心是一个准对称异源二聚体，为跨膜电子转移提供了两种可能的途径。过去的测量表明，在红光或近红外光的激发下，只有两种途径中的一种（a侧）被使用到任何显著程度。最近，伍德伯里博士发现，在200秒的激光脉冲中，蓝光激发到反应中心的Soret波段，沿着交替或b侧途径产生光化学反应。所形成的状态的光谱特征，以及它们在低温下的长（纳秒）寿命，都具有涉及b侧细菌内生菌素阴离子的电荷分离状态的特征。这为尝试理解反应中心b侧电子转移的机制和功能的一系列全新测量打开了大门。这个研究项目的中心是电荷分离态的自由能之间的相互作用，它可以通过改变P/P+势的突变体在很大范围内变化，以及用于激发的光子的能量。初步数据表明，b侧电子转移的能量越有利，启动b侧转移所需的光子能量越低。b侧电子转移的生理功能尚不清楚。一种可能性是，它可以快速地淬灭由反应中心辅因子和周围色氨酸残基吸收蓝光或紫外光形成的激发态。几个涉及破坏b侧转移的突变体的实验计划，看看反应中心是否更容易受到紫外线和蓝光的损害，而没有功能性b侧光化学。最后，为了产生一系列具有光谱定义功能特性的新突变体（损失或a侧电子转移，增加b侧转移等），Woodbury博士将开发一种定向进化方法，该方法完全利用光学手段从大型反应中心突变体库中筛选和选择细胞。

Light-initiated electron transfer within the photosynthetic reaction center is the primary event in solar energy conversion by photosynthetic organisms. The core of the photosynthetic reaction center from purple nonsulfur bacteria is a quasi-symmetric heterodimer, providing two potential pathways for transmembrane electron transfer. Past measurements have demonstrated that only one of the two pathways (the A-side) is used to any significant extent upon excitation with red or near infrared light. Recently, Dr. Woodbury has found that excitation with blue light into the Soret band of the reaction center gives rise to photochemistry along the alternate or B-side pathway apparently within a 200 fs laser pulse. The spectral signature of the states formed, and their long (nanosecond) lifetime at low temperature, are characteristic of charge separated states involving the anion of the B-side bacteriopheophytin. This opens the door to a whole new array of measurements attempting to understand both the mechanism and the function of B-side electron transfer in the reaction center. This research project centers around the interplay between the free energy of the charge separated states, which can be varied over a large range using mutants that alter the P/P+ potential, and the energy of the photon used in excitation. Preliminary data suggests that the more favorable the energetics of B-side electron transfer, the lower the energy of the photon that is required to initiate B-side transfer. The physiological function of B-side electron transfer is not yet known. One possibility is that it could rapidly quench the excited states formed from blue or UV light absorption by reaction center cofactors and surrounding tryptophan residues. Several experiments involving mutants that disrupt B-side transfer are planned to see if reaction centers are more susceptible to UV and blue light damage without functional B-side photochemistry. Finally, in order to generate a series of new mutants with spectrally defined functional properties (loss or A-side electron transfer, increased B-side transfer, etc.), Dr. Woodbury will develop a directed evolution methodology that utilizes entirely optical means of both screening and selecting cells from a large library of reaction center mutants.