SAXS STUDIES OF PROTEIN FOLDING INTERMEDIATES

我们建议在两个水平上研究蛋白质折叠中间体 (a)在合适的平衡条件下，我们可以稳定 蛋白质的部分折叠状态和（B）通过直接动力学 使用停流时间分辨SAXS进行测量。 小角X射线散射数据产额 的构象状态的总体大小和形状指示。 蛋白 细胞色素的高质量、高角度散射谱 c在不同的变性剂浓度下测量。 我们将 使用奇异值分解分析和 变性剂结合模型来确定平衡是否 中间状态存在。 该分析遵循用于 区分溶菌酶的折叠中间体（Chen，J. Mol. Chem. Biol. 261，658-671）。 A态，部分折叠的中间态 在低pH下加入盐得到的脱辅基肌红蛋白具有 用SAXS表征。 脱辅基肌红蛋白A状态稳定 TFA、TCA和KCl表现出不同程度的压实， 这表明并不存在唯一的A状态。

We propose to study protein folding intermediates at two levels (a) under suitable equilibrium conditions where we can stabilize partially folded states of proteins and (b) by direct kinetic measurements using stopped-flow time-resolved SAXS. SAXS data yield overall size and shape indication of the conformational state of a protein. High-quality, high-angle scattering profiles for cytochrome c have been measured at varying denaturant concentrations. We will use the combination of singular value decomposition analysis and a denaturant binding model to determine whether an equilibrium intermediate state exists. This analysis follows the method used to distinguish a folding intermediate for lysozyme (Chen, J. Mol. Biol. 261, 658-671). The A-states, partially folded intermediate states obtained at low pH with the addition of salt, of apomyoglobin have been characterized with SAXS. The apomyoglobin A-states stabilized with TFA, TCA, and KCl demonstrated different degrees of compaction, suggesting that there is not a unique A-state.