Chang Shanyan, Mizuno Misao, Ishikawa Haruto, Mizutani Yasuhisa
Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560-0043, Japan.
Phys Chem Chem Phys. 2018 Jan 31;20(5):3363-3372. doi: 10.1039/c7cp06287g.
Protein dynamics of human adult hemoglobin and its mutants restricted in R and T quaternary states following ligand photolysis were studied by time-resolved resonance Raman spectroscopy. In the time-resolved spectra, we observed spectral changes of in-plane stretching modes of heme and the iron-histidine stretching mode of the Fe-His bond for all the hemoglobin samples. The βD99N mutant, which adopts the R state in both the ligand-bound and the deoxy forms, showed similar temporal behaviors in time-resolved resonance Raman spectra as wild-type recombinant hemoglobin until 10 μs, consistent with the fact that the mutant undergoes only the tertiary structural changes in the R state. The βN102T mutant, which adopts the T state in both the ligand-bound and the deoxy forms, showed much slower tertiary structural changes, suggesting that the EF helical motion is decelerated by the change of the intersubunit interactions. The present data indicate that the allosteric kinetic response between the interhelical hydrogen bonds of the EF helices and the intersubunit hydrogen bonds is bidirectional. The implications of these results for understanding the allosteric pathway of Hb are discussed in detail.
通过时间分辨共振拉曼光谱研究了人类成人血红蛋白及其在配体光解后受限处于R态和T态四级结构的突变体的蛋白质动力学。在时间分辨光谱中，我们观察到了所有血红蛋白样品中血红素面内拉伸模式以及Fe-His键中铁-组氨酸拉伸模式的光谱变化。βD99N突变体在配体结合和脱氧形式下均采用R态，在10微秒之前，其时间分辨共振拉曼光谱中的时间行为与野生型重组血红蛋白相似，这与该突变体在R态仅发生三级结构变化的事实一致。βN102T突变体在配体结合和脱氧形式下均采用T态，其三级结构变化要慢得多，这表明EF螺旋运动因亚基间相互作用的改变而减速。目前的数据表明，EF螺旋间的螺旋间氢键与亚基间氢键之间的变构动力学响应是双向的。详细讨论了这些结果对理解血红蛋白变构途径的意义。
