Borgstahl G E, Williams D R, Getzoff E D
Department of Molecular Biology, Scripps Research Institute, La Jolla, California 92037, USA.
Biochemistry. 1995 May 16;34(19):6278-87. doi: 10.1021/bi00019a004.
A photosensing protein directs light energy captured by its chromophore into a photocycle. The protein's structure must accommodate the photocycle and promote the resulting chemical or conformational changes that lead to signal transduction. The 1.4 A crystallographic structure of photoactive yellow protein, determined by multiple isomorphous replacement methods, provides the first view at atomic resolution of a protein with a photocycle. The alpha/beta fold, which differs from the original chain tracing, shows striking similarity to distinct parts of the signal transduction proteins profilin and the SH2 domain. In the dark state structure of photoactive yellow protein, the novel 4-hydroxycinnamyl chromophore, covalently attached to Cys69, is buried within the major hydrophobic core of the protein and is tethered at both ends by hydrogen bonds. In the active site, the yellow anionic form of the chromophore is stabilized by hydrogen bonds from the side chains of Tyr42 and buried Glu46 to the phenolic oxygen atom and by electrostatic complementarity with the positively charged guanidinium group of Arg52. Thr50 further interlocks Tyr42, Glu46, and Arg52 through a network of active site hydrogen bonds. Arg52, located in a concavity of the protein surface adjacent to the dominant patch of negative electrostatic potential, shields the chromophore from solvent and is positioned to form a gateway for the phototactic signal. Overall, the high-resolution structure of photoactive yellow protein supports a mechanism whereby electrostatic interactions create an active site poised for photon-induced rearrangements and efficient protein-mediated signal transduction.
一种光感应蛋白将其发色团捕获的光能导入一个光循环中。该蛋白的结构必须适应光循环，并促进由此产生的导致信号转导的化学或构象变化。通过多同晶置换法确定的光活性黄色蛋白的1.4埃晶体结构，首次在原子分辨率下展示了具有光循环的蛋白。α/β折叠与最初的链追踪不同，与信号转导蛋白丝切蛋白和SH2结构域的不同部分有显著相似性。在光活性黄色蛋白的暗态结构中，共价连接到半胱氨酸69上的新型4-羟基肉桂基发色团被埋在蛋白的主要疏水核心内，并通过氢键在两端固定。在活性位点，发色团的黄色阴离子形式通过酪氨酸42和埋藏的谷氨酸46的侧链与酚氧原子之间的氢键以及与精氨酸52带正电的胍基的静电互补作用而稳定。苏氨酸50通过活性位点氢键网络进一步使酪氨酸42、谷氨酸46和精氨酸52相互连锁。精氨酸52位于蛋白表面与主要负静电势区域相邻的凹陷处，保护发色团免受溶剂影响，并定位形成光趋信号的通道。总体而言，光活性黄色蛋白的高分辨率结构支持一种机制，即静电相互作用产生一个为光子诱导的重排和高效蛋白介导的信号转导做好准备的活性位点。
