Department of Chemistry, University of Pennsylvania , Philadelphia, Pennsylvania 19104-6323, United States.
J Am Chem Soc. 2013 Sep 18;135(37):13914-26. doi: 10.1021/ja4067404. Epub 2013 Sep 5.
This work reports the first example of a single-chain protein computationally designed to contain four α-helical segments and fold to form a four-helix bundle encapsulating a supramolecular abiological chromophore that possesses exceptional nonlinear optical properties. The 109-residue protein, designated SCRPZ-1, binds and disperses an insoluble hyperpolarizable chromophore, ruthenium(II) [5-(4'-ethynyl-(2,2';6',2″-terpyridinyl))-10,20-bis(phenyl)porphinato]zinc(II)-(2,2';6',2″-terpyridine)(2+) (RuPZn) in aqueous buffer solution at a 1:1 stoichiometry. A 1:1 binding stoichiometry of the holoprotein is supported by electronic absorption and circular dichroism spectra, as well as equilibrium analytical ultracentrifugation and size exclusion chromatography. SCRPZ-1 readily dimerizes at micromolar concentrations, and an empirical redesign of the protein exterior produced a stable monomeric protein, SCRPZ-2, that also displayed a 1:1 protein:cofactor stoichiometry. For both proteins in aqueous buffer, the encapsulated cofactor displays photophysical properties resembling those exhibited by the dilute RuPZn cofactor in organic solvent: femtosecond, nanosecond, and microsecond time scale pump-probe transient absorption spectroscopic data evince intensely absorbing holoprotein excited states having large spectral bandwidth that penetrate deep in the near-infrared energy regime; the holoprotein electronically excited triplet state exhibits a microsecond time scale lifetime characteristic of the RuPZn chromophore. Hyper-Rayleigh light scattering measurements carried out at an incident irradiation wavelength of 1340 nm for these holoproteins demonstrate an exceptional dynamic hyperpolarizabilty (β1340 = 3100 × 10(-30) esu). X-ray reflectivity measurements establish that this de novo-designed hyperpolarizable protein can be covalently attached with high surface density to a silicon surface without loss of the cofactor, indicating that these assemblies provide a new approach to bioinspired materials that have unique electro-optic functionality.
这项工作报道了首例通过计算设计的单链蛋白，该蛋白包含四个α-螺旋片段，并能折叠形成四螺旋束，将超分子非生物生色团封装在内，该生色团具有出色的非线性光学性能。该 109 残基蛋白命名为 SCRPZ-1，可在水缓冲溶液中以 1:1 的化学计量比结合和分散不溶性可极化生色团[钌（II）[5-(4′-乙炔基-(2,2′;6′,2″-三联吡啶基))-10,20-二(苯基)卟啉基]锌(II)-(2,2′;6′,2″-三联吡啶)(2+)]（RuPZn）。电子吸收和圆二色性光谱以及平衡分析超速离心和尺寸排阻色谱均支持全蛋白的 1:1 结合化学计量比。SCRPZ-1 在微摩尔浓度下很容易二聚化，并且对蛋白外表面进行经验性重新设计产生了稳定的单体蛋白 SCRPZ-2，该蛋白也显示出 1:1 的蛋白辅因子化学计量比。对于水缓冲液中的两种蛋白，封装的辅因子显示出与在有机溶剂中稀释的 RuPZn 辅因子相似的光物理性质：飞秒、纳秒和微秒时间尺度的泵浦-探测瞬态吸收光谱数据表明，强烈吸收的全蛋白激发态具有大的光谱带宽，可穿透近红外能量区域；全蛋白电子激发三重态具有微秒时间尺度的寿命，这是 RuPZn 生色团的特征。这些全蛋白在 1340nm 入射辐照波长下进行的超瑞利光散射测量表明，其具有异常高的动态极化率（β1340=3100×10(-30)esu）。X 射线反射率测量表明，这种从头设计的可极化蛋白可以共价连接到硅表面上，而不会损失辅因子，这表明这些组装体为具有独特电光功能的仿生材料提供了一种新方法。
