光谱法研究尿素对水溶液中血红蛋白构象的影响

应用荧光猝灭法和动态光散射法测定尿素-水混合溶剂中血红蛋白(Hb)与联苯胺的结合距离和Hb的流体动力学半径. 结合Hb的荧光光谱和吸收光谱, 探讨尿素与蛋白质分子在水溶液中相互作用的机理及其对蛋白质构象的影响. 结果显示, 尿素分子取代水分子在蛋白质周围形成溶剂化层, 并与骨架肽链和亲水侧链形成氢键, 从而积聚在蛋白质分子表面. 尿素分子与蛋白质分子之间的直接相互作用对蛋白质的构象具有复杂的影响, 高浓度的尿素-水混合溶剂破坏蛋白质的构象, 而低浓度的混合溶剂则有利于蛋白质形成更紧密的构象. 在高浓度的尿素-水混合溶剂中, Hb血红素疏水空穴失去原有的三级结构后形成一个与熔球态相类似的结构.

Spectroscopic Studies on the Influence of Urea on the Conformation of Hemoglobin in Aqueous Solutions

The binding distance of benzidine to hemoglobin (Hb) and the hydrodynamic radius of Hb in urea-water mixtures were determined by fluorescence quenching and dynamic light scattering, respectively. These data, combined with fluorescence spectra and absorption spectra, were utilized to investigate the interaction between urea and protein as well as its influence on the conformation of protein in aqueous solutions. The results indicated that urea accumulated on the surface of the protein due to its ability to displace water molecules in the solvation shell and to form hydrogen-bond with peptides and hydrophilic side chains, which exhibited a complex influence on protein conformation. It was evident that urea-water mixtures at high urea concentration destabilized the protein conformation whereas those at low urea concentration promoted a more compact one. In the mixtures at high urea concentration, the heme cavity of Hb was found to be of unfolded structure, however similar to the molten globubar state.