Hulsker Rinske, Baranova Maria V, Bullerjahn George S, Ubbink Marcellus
Leiden Institute of Chemistry, Leiden University, Gorlaeus Laboratories, P.O. Box 9502, 2300 RA Leiden, The Netherlands.
J Am Chem Soc. 2008 Feb 13;130(6):1985-91. doi: 10.1021/ja077453p. Epub 2008 Jan 18.
The nature of transient protein complexes can range from a highly dynamic ensemble of orientations to a single well-defined state. This represents variation in the equilibrium between the encounter and final, functional state. The transient complex between plastocyanin (Pc) and cytochrome f (cytf) of the cyanobacterium Prochlorothrix hollandica was characterized by NMR spectroscopy. Intermolecular pseudocontact shifts and chemical shift perturbations were used as restraints in docking calculations to determine the structure of the wild-type Pc-cytf complex. The orientation of Pc is similar to orientations found in Pc-cytf complexes from other sources. Electrostatics seems to play a modest role in complex formation. A large variability in the ensemble of lowest energy structures indicates a dynamic nature of the complex. Two unusual hydrophobic patch residues in Pc have been mutated to the residues found in other plastocyanins (Y12G/P14L). The binding constants are similar for the complexes of cytf with wild-type Pc and mutant Pc, but the chemical shift perturbations are smaller for the complex with mutant Pc. Docking calculations for the Y12G/P14L Pc-cytf complex did not produce a converged ensemble of structures. Simulations of the dynamics were performed using the observed averaged NMR parameters as input. The results indicate a surprisingly large amplitude of mobility of Y12G/P14L Pc within the complex. It is concluded that the double mutation shifts the complex further from the well-defined toward the encounter state.
瞬时蛋白质复合物的性质范围可以从高度动态的取向集合到单一明确的状态。这代表了相遇状态与最终功能状态之间平衡的变化。通过核磁共振光谱对荷兰原绿球藻的质体蓝素（Pc）和细胞色素f（cytf）之间的瞬时复合物进行了表征。分子间的赝接触位移和化学位移扰动被用作对接计算中的约束条件，以确定野生型Pc-cytf复合物的结构。Pc的取向与其他来源的Pc-cytf复合物中的取向相似。静电作用在复合物形成中似乎起适度作用。最低能量结构集合中的较大变异性表明该复合物具有动态性质。Pc中的两个不寻常的疏水补丁残基已被突变为其他质体蓝素中发现的残基（Y12G/P14L）。cytf与野生型Pc和突变型Pc的复合物的结合常数相似，但与突变型Pc的复合物的化学位移扰动较小。Y12G/P14L Pc-cytf复合物的对接计算未产生收敛的结构集合。使用观察到的平均核磁共振参数作为输入进行动力学模拟。结果表明，复合物中Y12G/P14L Pc的迁移率具有惊人的大幅度。得出的结论是，双重突变使复合物进一步从明确状态向相遇状态转变。
