Giachini Lisa, Francia Francesco, Mallardi Antonia, Palazzo Gerardo, Carpenè Emilio, Boscherini Federico, Venturoli Giovanni
Dipartimento di Fisica, Università di Bologna, Bologna, Italy.
Biophys J. 2005 Mar;88(3):2038-46. doi: 10.1529/biophysj.104.050971. Epub 2004 Dec 21.
Binding of transition metal ions to the reaction center (RC) protein of the photosynthetic bacterium Rhodobacter sphaeroides has been previously shown to slow light-induced electron and proton transfer to the secondary quinone acceptor molecule, Q(B). On the basis of x-ray diffraction at 2.5 angstroms resolution a site, formed by AspH124, HisH126, and HisH128, has been identified at the protein surface which binds Cd(2+) or Zn(2+). Using Zn K-edge x-ray absorption fine structure spectroscopy we report here on the local structure of Zn(2+) ions bound to purified RC complexes embedded into polyvinyl alcohol films. X-ray absorption fine structure data were analyzed by combining ab initio simulations and multiparameter fitting; structural contributions up to the fourth coordination shell and multiple scattering paths (involving three atoms) have been included. Results for complexes characterized by a Zn to RC stoichiometry close to one indicate that Zn(2+) binds two O and two N atoms in the first coordination shell. Higher shell contributions are consistent with a binding cluster formed by two His, one Asp residue, and a water molecule. Analysis of complexes characterized by approximately 2 Zn ions per RC reveals a second structurally distinct binding site, involving one O and three N atoms, not belonging to a His residue. The local structure obtained for the higher affinity site nicely fits the coordination geometry proposed on the basis of x-ray diffraction data, but detects a significant contraction of the first shell. Two possible locations of the second new binding site at the cytoplasmic surface of the RC are proposed.
过渡金属离子与光合细菌球形红杆菌的反应中心（RC）蛋白的结合，先前已表明会减缓光诱导的电子和质子向次级醌受体分子Q(B)的转移。基于2.5埃分辨率的X射线衍射，已在蛋白质表面鉴定出一个由AspH124、HisH126和HisH128形成的位点，该位点可结合Cd(2+)或Zn(2+)。我们在此使用Zn K边X射线吸收精细结构光谱，报告嵌入聚乙烯醇薄膜中的纯化RC复合物结合的Zn(2+)离子的局部结构。通过结合从头算模拟和多参数拟合对X射线吸收精细结构数据进行分析；已纳入直至第四配位层的结构贡献和多散射路径（涉及三个原子）。以接近1的Zn与RC化学计量比为特征的复合物结果表明，Zn(2+)在第一配位层中结合两个O原子和两个N原子。更高层的贡献与由两个His、一个Asp残基和一个水分子形成的结合簇一致。对每个RC约有2个Zn离子的复合物的分析揭示了第二个结构上不同的结合位点，涉及一个O原子和三个N原子，不属于His残基。从高亲和力位点获得的局部结构很好地符合基于X射线衍射数据提出的配位几何，但检测到第一配位层有明显收缩。提出了RC细胞质表面第二个新结合位点的两个可能位置。
