SMALL ANGLE X-RAY SCATTERING STUDY OF SOLVENT ISOTOPE EFFECT AND PROTEIN STABILI

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 我们提出了小角X射线散射实验（SAXS）在SSRL研究的影响，H2O-D2 O溶剂取代效应的球状蛋白质的结构稳定性。蛋白质自然存在于水溶液中。然而，通常使用各种实验技术将蛋白质溶解在重水中用于生物物理研究，例如，红外光谱、拉曼散射、核磁共振和中子散射，以避免在存在H2O的情况下测量蛋白质所涉及的困难。为了有效，这些研究假设氘同位素取代不会显著影响生物分子结构或动力学，基于D2 O是引起最温和可能扰动的溶剂的想法。然而，实验数据，特别是热力学研究表明，溶剂取代会对蛋白质在溶液中的稳定性产生显着影响。特别地，D2 O取代H2O使蛋白质稳定以抵抗热变性和尿素诱导的变性。更重要的是，已经发现，一些蛋白质似乎在D2 O和H2O中采用不同的结构构象，而其他溶液条件相同。小角散射实验是唯一能直接提供溶液中蛋白质分子整体结构构象信息的技术。这将极大地补充热力学结果，并有助于我们理解蛋白质变性的氘同位素效应。对于这个拟议的项目，我们将监测两个模型蛋白质的结构变化，母鸡？分别以鸡蛋溶菌酶和牛血清白蛋白（BSA）为研究对象，研究了D2 O和H2O对BSA热变性和尿素诱导变性的影响。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. We propose small angle x-ray scattering experiments (SAXS) at SSRL to study the influence of H2O-D2O solvent substitution effect on the structural stability of globular proteins. Proteins naturally exist in aqueous solutions. However, it is common to dissolve proteins in heavy water for biophysical studies using various experimental techniques, e.g., infrared spectroscopy, Raman scattering, nuclear magnetic resonance, and neutron scattering, in order to avoid the difficulties involved in the measurement of proteins with the presence of H2O. To be valid, these studies assume that deuterium isotopic substitution does not influence biomolecular structure or dynamics significantly, based on the idea that D2O is the solvent that causes the mildest possible perturbation. However, experimental data, especially thermodynamic studies, have shown that solvent substitution can cause marked effects on the stability of proteins in solution. In particular, the substitution of D2O for H2O stabilizes the proteins against thermal denaturation and urea-induced denaturation. More significantly, it has been found that some proteins appear to adopt different structural conformations in D2O and H2O, with other solution conditions being identical. The small angle scattering experiment is the only technique that directly provides essential information on the global structural conformation of protein molecules in solution. It will greatly complement the thermodynamic results and contribute significantly to our understanding of deuterium isotopic effects on protein denaturation. For this proposed project, we will monitor the structural change of two model proteins, hen?s egg lysozyme and bovine serum albumin (BSA) dissolved in H2O and D2O, respectively, in the thermal denaturation and urea-induced denaturation processes, and study the influence on the denaturation behavior by the substitution of H2O with D2O.