Herskovits T T, Ibanez V S
Biochemistry. 1976 Dec 28;15(26):5715-21. doi: 10.1021/bi00671a006.
The subunit dissociation of human hemoglobin A by the aliphatic acid salts at neutral pH has been investigated by light-scattering molecular-weight measurements at 630 nm. Dissociation of hemoglobin tetramers to alphabeta dimers is observed in essentially all experiments at low to intermediate levels of salt concentrations, below the denaturation transitions, described in the accompanying paper (Ibanez, V.S., and Herskovits, T.T. (1976), Biochemistry 15, preceding paper in this issue). The effectiveness of the salts as subunit dissociating agent, reflected by the slopes, s, of the plots of deltaGDdegrees, the standard free energy of dissociation, vs. [D], the salt concentration, is found to increase with increasing alkyl chain length or hydrocarbon content of the salt. Estimates of the apparent number of amino acid sites at the areas of contact per alphabeta dimer formed, N', based on the slopes of the higher members of the series have been obtained using the equation, deltaGDdegrees = deltaGD,Wdegrees - 2N'RTKB[D]. Independent estimates of the binding constant, KB, required for these calculations were based on free-energy transfer data of hydrophobic amino acid alkyl groups and protein denaturation data. Our estimates of N' denaturation data. Our estimates of N' obtained with the more reliable data of the higher members of salt series are in the ranges of 19 and 27 amino acid groups, shown by the x-ray crystallographic structure of horse and human hemoglobin of Perutz (Perutz, M.F., et al. (1968), Nature (London) 219, 131) and Fermi ((1975) J. Mol. Biol. 97, 237) for the smaller alpha1 beta2 contact areas in the tetrameric structure. The lower estimates than 27 based on our dissociation of human hemoglobin suggest that several of the amino acid residues in the contact areas of the subunits are partially exposed to solvent. The increasing effectiveness of the higher mn imporant source of stabilization of the tetrameric structure of hemoglobin.
在中性pH条件下，通过在630nm处进行光散射分子量测量，研究了脂肪酸盐对人血红蛋白A的亚基解离作用。在低至中等盐浓度水平（低于随附论文（伊瓦涅斯，V.S.，和赫斯科维茨，T.T.（1976年），《生物化学》15卷，本期前文）中描述的变性转变）的基本上所有实验中，均观察到血红蛋白四聚体解离为αβ二聚体。盐作为亚基解离剂的有效性，由解离标准自由能ΔG°D与盐浓度[D]的关系图的斜率s反映，发现其随着盐的烷基链长度或烃含量的增加而增加。基于该系列中较高成员的斜率，使用方程ΔG°D = ΔG°D,W - 2N'RTKB[D]，获得了每个形成的αβ二聚体接触区域处氨基酸位点的表观数量N'的估计值。这些计算所需的结合常数KB的独立估计值基于疏水氨基酸烷基的自由能转移数据和蛋白质变性数据。我们对N'变性数据的估计。我们使用盐系列中较高成员的更可靠数据获得的N'估计值在19至27个氨基酸基团的范围内，这与佩鲁茨（佩鲁茨，M.F.等人（1968年），《自然》（伦敦）219卷，131页）和费米（（1975年）《分子生物学杂志》97卷，237页）报道的马和人血红蛋白的x射线晶体结构中四聚体结构较小的α1β2接触区域相符。基于我们对人血红蛋白解离的结果，低于27的较低估计值表明亚基接触区域中的几个氨基酸残基部分暴露于溶剂中。较高成员的有效性增加是血红蛋白四聚体结构稳定的重要来源。
