Measurement of biomolecular interactions and association via light scattering

通过光散射测量生物分子相互作用和缔合

• 批准号: 8741359
• 负责人: Allen P Minton
• 金额: $ 20.29万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8741359
• 关键词: Accounting; Amines; Data; Diffusion; Dissociation; Equilibrium; Exclusion; Guanidinium Chloride; Guanine Nucleotides; Guanosine Triphosphate; Laboratories; Light; Measurement; Modeling; Molecular Conformation; Molecular Weight; Monitor; Oxides; Phase Transition; Potassium; Proteins; Solutions; Techniques; Thermodynamics; analog; light scattering; macromolecule; protein function; stoichiometry

1. The effect of guanidine hydrochloride (GuHCl) and trimethyl-N-amine oxide (TMAO) upon the equilibrium dissociation of tetrameric to dimeric cyanmethemoglobin (CNmetHb) was studied by measurement of dynamic light scattering. This measurement yields the intensity average diffusion coefficient of the protein, which increases as the protein dissociates. The results showed that the increasing tendency of CNmetHb to dissociate with added GuHCl may be inhibited by addition of sufficient amounts of TMAO. The stabilizing effect of TMAO is attributed largely to volume exclusion (D. Wu). 2. The self-association of the bacterial septation protein FtsZ in the presence of GTP or a GTP analogue, GMPCPP, and 500 mM potassium was studied as a function of protein and Mg concentration using single-angle and multi-angle static light scattering. The results reveal that when the concentration of protein exceeds a Mg-dependent critical value, FtsZ undergoes a concerted formation, akin to a second-order phase transition to a narrow distribution of high molecular weight oligomers with a stoichiometry of ca 100 (GTP) or 160 (GMPCPP). Above the transition concentration, the amount, but not the size, of oligomer increases, while that of low molecular weight species remains roughly constant (Monterroso, Rivas). All of the data obtained from the several types of measurements may be accounted for semiquantitatively in the context of a model, according to which the stable oligomer is a cyclic species, the size of which reflects the tendency of the protein fibrils to flex or curve in solution in the presence of different concentrations of Mg and the particular guanine nucleotide present (B. Monterroso, G. Rivas, CIB).

1.用动态光散射法研究了盐酸胍和三甲基N-胺氧化物对四聚体平衡解离成二聚体氰化高铁血红蛋白的影响。这种测量产生了蛋白质的强度平均扩散系数，该系数随着蛋白质的解离而增加。结果表明，加入足量的TMAO可以抑制CNmetHb随盐酸葡胺浓度的增加而升高的趋势。TMAO的稳定作用很大程度上归因于体积排斥(D.Wu)。 2.用单角和多角静态光散射法研究了细菌分离蛋白FtsZ在GTP或GTP类似物GMPCPP和500 mM钾存在下的自结合作用，并研究了其与蛋白质和镁浓度的关系。结果表明，当蛋白质浓度超过一个依赖于镁的临界值时，FtsZ发生了协同形成，类似于二级相变，形成了化学计量比约为100(GTP)或160(GMPCPP)的窄分布的高相对分子质量齐聚物。在转变浓度以上，齐聚物的数量增加，但大小不变，而低分子量物种的数量大致保持不变(蒙特罗索，里瓦斯)。从几种类型的测量获得的所有数据可以在一个模型的背景下半定量地解释，根据该模型，稳定的低聚物是环状物种，其大小反映了在存在不同浓度的镁和存在的特定鸟嘌呤核苷酸的情况下蛋白质原纤维在溶液中弯曲或弯曲的趋势(B.蒙特罗索，G.Rivas，CIB)。