Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan 609-735, Korea.
J Phys Chem B. 2012 May 24;116(20):5819-30. doi: 10.1021/jp300176q. Epub 2012 May 11.
The effect of the solvent viscosity on the dynamics of NO rebinding to myoglobin (Mb) and hemoglobin (Hb) was examined by femtosecond (fs) time-resolved vibrational spectroscopy after photodeligation of NO from MbNO and HbNO in various viscous solutions at 283 K using a 580 nm excitation pulse. The rebinding kinetics of NO to both Mb and Hb were nonexponential, but their dependence on the solvent viscosity was different. The rate of NO rebinding to Mb increased with increasing solution viscosity, which was achieved by increasing the glycerol content in glycerol/water mixture. In contrast, the rate of NO rebinding to Hb was independent of the solution viscosity but faster than the fastest rate of NO rebinding observed in Mb. The dynamics of conformational relaxation of the protein after deligation were also measured by probing the evolution of the amide band. The effect of the solvent viscosity on the kinetics of conformational relaxation in both proteins was also quite different. The conformational relaxation of Mb became slower with increasing solution viscosity. On the other hand, the conformational relaxation of Hb was independent of the solution viscosity but slower than the slowest kinetics of Mb. The inverse correlation in the kinetics of conformational relaxation and NO rebinding suggests that the barrier of NO rebinding increases as the conformation of the protein relaxes toward the deligated structure after NO dissociation. The rebinding kinetics of NO to both proteins was well described by a kinetic model incorporating a time-dependent barrier for rebinding and exponential translocations between three states for dissociated NO.
采用飞秒时间分辨振动光谱技术，在 283 K 下，用 580nm 的激发脉冲光解 MbNO 和 HbNO 于不同粘性溶液中，考察了溶剂粘度对 NO 重新结合到肌红蛋白（Mb）和血红蛋白（Hb）的动力学的影响。NO 重新结合到 Mb 和 Hb 的反应动力学均非指数型，但它们对溶剂粘度的依赖性不同。NO 重新结合到 Mb 的速率随溶液粘度的增加而增加，这是通过增加甘油/水溶液中甘油的含量实现的。相比之下，NO 重新结合到 Hb 的速率与溶液粘度无关，但比在 Mb 中观察到的最快的 NO 重新结合速率快。通过探测酰胺带的演化，还测量了配体光解后蛋白质构象弛豫的动力学。溶剂粘度对两种蛋白质构象弛豫动力学的影响也大不相同。Mb 的构象弛豫随溶液粘度的增加而变慢。另一方面，Hb 的构象弛豫与溶液粘度无关，但比 Mb 的最慢动力学慢。构象弛豫和 NO 重新结合动力学的反相关表明，随着 NO 离解后蛋白质构象弛豫到去配体结构，NO 重新结合的势垒增加。NO 重新结合到两种蛋白质的动力学很好地用一个动力学模型描述，该模型包含了重新结合的时变势垒和离解态 NO 之间的三个状态的指数型跃迁。
