Santiago Patricia S, Moreira Leonardo M, de Almeida Erika V, Tabak Marcel
Instituto de Quimica de São Carlos, Universidade de São Paulo, 13560-970 São Carlos, SP, Brazil.
Biochim Biophys Acta. 2007 Apr;1770(4):506-17. doi: 10.1016/j.bbagen.2006.11.005. Epub 2006 Nov 23.
The effects of two ionic surfactants on the oligomeric structure of the giant extracellular hemoglobin of Glossoscolex paulistus (HbGp) in the oxy - form have been studied through the use of several spectroscopic techniques such as electronic optical absorption, fluorescence emission, light scattering, and circular dichroism. The use of anionic sodium dodecyl sulphate (SDS) and cationic cethyltrimethyl ammonium chloride (CTAC) has allowed to differentiate the effects of opposite headgroup charges on the oligomeric structure dissociation and hemoglobin autoxidation. At pH 7.0, both surfactants induce the protein dissociation and a significant oxidation. Spectral changes occur at very low CTAC concentrations suggesting a significant electrostatic contribution to the protein-surfactant interaction. At low protein concentration, 0.08 mg/ml, some light scattering within a narrow CTAC concentration range occurs due to protein-surfactant precipitation. Light scattering experiments showed the dissociation of the oligomeric structure by SDS and CTAC, and the effect of precipitation induced by CTAC. At higher protein concentrations, 3.0 mg/ml, a precipitation was observed due to the intense charge neutralization upon formation of ion pair in the protein-surfactant precipitate. The spectral changes are spread over a much wider SDS concentration range, implying a smaller electrostatic contribution to the protein-surfactant interactions. The observed effects are consistent with the acid isoelectric point (pI) of this class of hemoglobins, which favors the intense interaction of HbGp with the cationic surfactant due to the existence of excess acid anionic residues at the protein surface. Protein secondary structure changes are significant for CTAC at low concentrations while they occur at significantly higher concentrations for SDS. In summary, the cationic surfactant seems to interact more strongly with the protein producing more dramatic spectral changes as compared to the anionic one. This is opposite as observed for several other hemoproteins. The surfactants at low concentrations produce the oligomeric dissociation, which facilitates the iron oxidation, an important factor modulating further oligomeric protein dissociation.
通过使用多种光谱技术，如电子吸收光谱、荧光发射光谱、光散射光谱和圆二色光谱，研究了两种离子表面活性剂对保罗氏光尾蚓（Glossoscolex paulistus）氧合形式的巨型细胞外血红蛋白（HbGp）寡聚结构的影响。使用阴离子十二烷基硫酸钠（SDS）和阳离子十六烷基三甲基氯化铵（CTAC）能够区分相反头基电荷对寡聚结构解离和血红蛋白自氧化的影响。在pH 7.0时，两种表面活性剂均诱导蛋白质解离并发生显著氧化。在非常低的CTAC浓度下就出现光谱变化，表明静电作用对蛋白质-表面活性剂相互作用有显著贡献。在低蛋白浓度（0.08 mg/ml）下，由于蛋白质-表面活性剂沉淀，在狭窄的CTAC浓度范围内会出现一些光散射。光散射实验表明SDS和CTAC均可使寡聚结构解离，以及CTAC诱导的沉淀效应。在较高蛋白浓度（3.0 mg/ml）下，由于蛋白质-表面活性剂沉淀中离子对形成时的强烈电荷中和作用，观察到沉淀现象。光谱变化分布在更宽的SDS浓度范围内，这意味着静电作用对蛋白质-表面活性剂相互作用的贡献较小。观察到的效应与这类血红蛋白的酸性等电点（pI）一致，由于蛋白质表面存在过量的酸性阴离子残基，这有利于HbGp与阳离子表面活性剂的强烈相互作用。低浓度时CTAC会引起蛋白质二级结构显著变化，而SDS则在显著更高的浓度下才会引起这种变化。总之，与阴离子表面活性剂相比，阳离子表面活性剂似乎与蛋白质的相互作用更强，产生更显著的光谱变化。这与其他几种血红蛋白的情况相反。低浓度的表面活性剂会导致寡聚体解离，这有利于铁的氧化，而铁氧化是调节进一步的寡聚蛋白质解离的一个重要因素。
