TIME-RESOLVED SAXS/WAXS OF PHOTOACTIVE YELLOW PROTEIN (PYP)

光活性黄色蛋白 (PYP) 的时间分辨萨克斯/蜡

• 批准号: 8168650
• 负责人: IHEE HYOTCHERL
• 金额: $ 1.08万
• 依托单位: ILLINOIS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8168650
• 关键词: 4-coumaric acid; Computer Retrieval of Information on Scientific Projects Database; Crowding; Excision; Funding; Furuncles; Glues; Grant; Hour; Hydrogen Bonding; Hydrophobic Interactions; Hydroxylamine; Institution; Link; Pattern; Phase; Process; Proteins; Reaction; Research; Research Personnel; Resources; Roentgen Rays; Shapes; Solutions; Source; System; Time; United States National Institutes of Health; chromophore; covalent bond; thioester

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. A well-developed light-responsive system, photoactive yellow protein (PYP), contains p- coumaric acid as the chromophore, which is covalently linked to the Cys69 residue by a thioester bond (1) and has a maximum absorbance at 446 nm (2). The chromophore interacts with nearby residues through hydrogen bonds and hydrophobic interactions, and this hydrogen bond networking acts as the glue which makes holo-PYP (PYP with the chromophore) more compact and stable than apo-PYP (PYP without the chromophore). Therefore removal of the chromophore from holo-PYP induces unfolding process into apo-PYP and provides a unique opportunity to study protein unfolding process. Due to the covalent linking to PYP, the chromophore is not removed by denaturing or boiling, but can be irreversibly detached by reacting with hydroxylamine, which specifically reacts with the chromophore covalent bond The reaction of hydroxylamine with holo-PYP to produce apo-PYP occurs with a slow rate constant of a few hours. X-ray solution scattering is useful for studying protein unfolding process in solution phase because it provides information concerning the approximate size, shape, and conformational changes of protein. A combination of both small-angle (SAXS) and wide-angle X-ray solution scattering (WAXS) will be used to study overall changes of shape and size by SAXS and conformational changes of protein by WAXS during the unfolding process. We will also investigate the effects of concentration on SAXS and WAXS patterns for both holo-PYP and apo-PYP to check whether the degree of crowding depends on the presence of the chromophore (4, 5).

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 发达的光响应性系统，光活性黄蛋白（PYP）包含p-丙甲酸作为发色团，它通过硫酯键（1）共价链接与Cys69残基（1），并在446 nm（2）时具有最大吸收率。发色团通过氢键和疏水相互作用与附近残基相互作用，而这种氢键网络充当胶水，使Holo-PYP（带有发色团的PYP）比Apo-PYP（无编成生园的PYP）更紧凑，更稳定。因此，从整体型中将发色团去除将展开的过程诱导到apo-pyp中，并为研究蛋白质展开过程提供了独特的机会。由于与PYP的共价相关，因此不能通过贬低或煮沸来去除染色体，但可以通过与羟胺反应来不可逆地脱离，该反应与染色体共键键特别反应，使羟胺与Holo-pyp的反应与Holo-pyp的反应产生apo-pyp，以使Apo-pyp与慢速速率持续不断变化。 X射线溶液散射可用于研究溶液相中的蛋白质展开过程，因为它提供了有关蛋白质的大小，形状和构象变化的信息。小角度（SAX）和广角X射线溶液散射（蜡）的组合将用于研究在展开过程中，萨克斯的形状和大小的总体变化以及蜡对蛋白质的构象变化。我们还将研究Holo-PYP和APO-PYP的浓度对SAX和蜡模式的影响，以检查拥挤程度是否取决于发色团的存在（4，5）。