Measurement of biomolecular association via static and dynamic light scattering

1.我们同时收集了非相互作用和相互作用蛋白质混合物的组合物依赖的静态和动态光散射数据，并开发了一个全球性的分析这两个数量的组合物依赖性。 与预期相反，添加动态光散射数据并没有提高仅使用静态光散射数据可获得的表征的分辨率。 这项工作的成果已提交出版。 （B.（roso） 2.我们已经构建了两种不同的仪器，旨在同时测量随时间变化的静态和动态光散射的蛋白质聚集。 第一种仪器是基于比色皿的仪器，将测量90 °静态和动态光散射。 第二种是一种仪器，它将自动从反应容器中提取聚集蛋白质的等分试样，并将其引入流动池中，用于测量多个角度的静态光散射和单个角度的动态散射。 我们的目标是获得，在一个单一的实验中，重均分子量的时间依赖性，Z-平均回转半径，和强度加权分布的扩散系数。 这三个量的建模应该提供一个高分辨率的图片的动力学计划的聚集。（A.阿特里角（费尔南德斯） 3.我们通过测量静态光散射的浓度依赖性，表征了胰岛素在宽pH值范围内的自缔合平衡（A. Attri）。 在pH 1.6时，胰岛素在醋酸盐缓冲液中为非缔合单体，在HCl中为极弱的自缔合二聚体。 在pH值为3和8之间，散射的浓度依赖性定量占一个简单的等键不定缔合计划。 在pH值为10时，散射的浓度依赖性是定量占由一个修改的等键计划，其中的平衡缔合常数为单体单体的添加是约5倍小于平衡缔合常数为单体添加到所有更高的低聚物。（A. Attri） 4.我们已经表征了tau蛋白和7 K分子量级分的肝素（一种具有高负电荷密度的硫酸化多糖）之间的相互作用。 散射强度的组成依赖性很好地描述了一个简单的1：1的异缔合。 这是首次将组成梯度-静态光散射应用于两种不同类型大分子之间的缔合。

1. We have simultaneously collected composition-dependent static and dynamic light scattering data on mixtures of non-interacting and interacting proteins, and developed a global analysis of the composition-dependence of both quantities. Contrary to expectation, addition of dynamic light scattering data did not enhance the resolution of characterization obtainable using static light scattering data only. Results of this work have been submitted for publication. (B. Monterroso) 2. We have constructed two different instruments designed to simultaneously measure time-dependent static and dynamic light scattering of proteins undergoing aggregation. The first instrument is a cuvette-based instrument that will measure 90o static and dynamic light scattering. The second is an instrument that will automatically extract aliquots of aggregating protein from a reaction vessel and introduce it into a flow cell for measurement of static light scattering at multiple angles and dynamic scattering at a single angle. Our goal is to obtain, in a single experiment, the time dependence of weight-average molecular weight, z-average radius of gyration, and the intensity-weighted distribution of diffusion coefficients. Modeling of the three quantities should provide a high resolution picture of the kinetic scheme of aggregation. (A. Attri, C. Fernandez) 3. We have characterized the self-association equilibria of insulin over a wide range of pH values, via measurement of the concentration dependence of static light scattering (A. Attri). At pH 1.6, insulin is a non-associating monomer in acetate buffer and a very weakly self-associating dimer in HCl. At pH values between 3 and 8, the concentration dependence of scattering is quantitatively accounted for by a simple isodesmic indefinite association scheme. At pH 10, the concentration dependence of scattering is quantitatively accounted for by a modified isodesmic scheme in which the equilibrium association constant for addition of monomer to monomer is about five times smaller than the equilibrium association constant for addition of monomer to all higher oligomers. (A. Attri) 4. We have characterized the interaction between tau protein and a 7K molecular weight fraction of heparin, a sulfated polysaccharide with high negative charge density. The composition dependence of the scattering intensity is well described by a simple 1:1 heteroassociation. This is the first application of composition gradient - static light scattering to the association between two different types of macromolecule.