Noncovalent Intermolecular Interactions In Biochemistry

基于光散射的可逆大分子缔合物的快速检测和定量表征方法是去年首次报道的。的年度报告已经扩展到能够检测和表征溶液中两种蛋白质之间的关联。我们已经证明了该技术的能力，以表征两种蛋白质的多个复合物，并表征在同一溶液中发生的自缔合和异缔合。最后，该技术已被用来表征的浓度依赖性分布的多个物种在一个单一的蛋白质的溶液中进行自我关联，形成线性低聚物的不定长度。 与Peter McPhie（NIDDK）合作，通过圆二色光谱研究了大分子拥挤对脱辅基肌红蛋白在热和冷变性方面的稳定性的影响。在pH值为2时，脱辅基肌红蛋白几乎完全变性，但在存在的三氯乙酸在毫摩尔浓度，采用紧凑的非原生结构称为？熔融球？其在低温（冰点以上）以及高温下展开。在足够高浓度的非离子多糖（葡聚糖T10）的存在下，50%热展开的温度升高，50%冷展开的温度降低，根据早期的预测排除体积理论。三氯乙酸和右旋糖酐对脱辅基肌红蛋白的温度依赖性椭圆率的影响可以用两态模型半定量地解释。

The light scattering based method for rapid detection and quantitative characterization of reversible macromolecular associations first reported in last year?s annual report has been extended to enable the detection and characterization of associations between two proteins in solution. We have demonstrated the capability of the technique to characterize multiple complexes of two proteins, and to characterize both self-association and hetero-association occurring in the same solution. Finally, the technique has been used to characterize the concentration-dependent distribution of multiple species in a solution of a single protein undergoing self-association to form linear oligomers of indefinite length. In collaboration with Peter McPhie (NIDDK), the effects of macromolecular crowding upon the stability of apomyoglobin with respect to denaturation by both heat and cold have been studied via circular dichroism spectroscopy. At pH 2, apomyoglobin is almost completely denatured, but in the presence of trichloroacetate at millimolar concentrations, adopts a compact non-native structure termed ?molten globule?, which unfolds at low temperatures (above freezing) as well as at high temperatures. In the presence of sufficiently high concentrations of a nonionic polysaccharide (dextran T10) the temperature for 50% heat unfolding is elevated and that for 50% cold unfolding is depressed, in accordance with earlier predictions of excluded volume theory. The effect of both trichloroacetate and dextran upon the temperature-dependent ellipticity of apomyoglobin may be accounted for semiquantitatively by a two-state model.