Properties of concentrated macromolecular solutions

1. 使用本实验室开发的技术和仪器测量了总浓度高达100 g/l的三种蛋白质（牛血清白蛋白、卵白蛋白和卵黏液样蛋白）和小分子渗透物三甲胺- n -氧化物（TMAO）的混合物的光散射强度的组成依赖性，并在以前的报告中进行了描述。结果是用这个实验室开发的适合于非缔合溶质在高浓度下表现出排斥性自相互作用和异质相互作用的二元混合物的形式分析的。结果可以用半经验模型来定量解释，其中蛋白质被表示为一个球体，其有效体积取决于它是与自身相互作用还是与TMAO相互作用，TMAO被表示为一个较小的恒定体积的球体。该模型的基本原理是，在测量条件下，这里研究的每个蛋白质都带一个净负电荷。因此，蛋白质分子之间的自相互作用具有排斥静电相互作用的元素，这在蛋白质分子与中性氧化三甲胺分子之间的相互作用中基本上是不存在的。

1. The composition dependence of the light scattering intensity of mixtures of each of three proteins, BSA, ovalbumin, and ovomucoid, and a small molecule osmolyte, trimethylamine-N-oxide (TMAO) at total concentrations of up to 100 g/l was measured using a technique and instrumentation developed in this laboratory and described in previous reports. The results were analyzed using formalism developed in this laboratory appropriate for binary mixtures of non-associating solutes exhibiting repulsive self- and hetero-interaction at high concentration. The results could be quantitatively accounted for by a semiempirical model in which the protein was represented as a sphere, the effective volume of which depended upon whether it was interacting with itself or with TMAO, and TMAO was represented as a smaller sphere of constant volume. The rationale for this model is that at under the conditions of measurement, each of the proteins studied here bears a net negative charge. Thus self-interactions between protein molecules have an element of repulsive electrostatic interaction that is largely absent from the interaction between a protein molecule and a neutral TMAO molecule. 2. The composition and temperature dependence of the sedimentation equilibrium of mixtures of each of three dilute tracer proteins (BSA, ovalbumin, and superoxide dismutase or SOD) and and two concentrated proteins (BSA and ovalbumin) at total concentrations of up to 100 g/l and temperatures between 5 and 37 degrees Centigrade, by means of a technique (nonideal tracer sedimentation equilibrium) and instrumentation developed in this laboratory, described in previous reports. The results could be quantitatively accounted for by semiempirical models, according to which each protein was represented as a sphere, self-interaction of concentrated protein was purely repulsive, and hetero-interaction of concentrated and dilute protein was primarily repulsive, but the consequences of repulsion reduced slightly due to a small attractive interaction that could be characterized as extremely weak heteroassociation (Kd of order 10 mM). The repulsive interactions were found to be independent of to within experimental uncertainty, and only one attractive interaction, that between tracer SOD and ovomucoid, exhibited a small but significant temperature dependence over the range studied. 3. In collaboration with German Rivas (CIB, Madrid) a mini-review on the subject of sedimentation equilibrium in highly nonideal solutions was written. The review describes experimental methods and general theory for quantitative interpretation of data.