CRYSTALS OF 143 KDA BOVINE INTERPHOTORECEPTOR RETINOID BINDING PRO

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 143 kDa的人/哺乳动物光感受器间类视黄酸结合蛋白(IRBP)与其他类视黄酸结合蛋白没有同源性，这是不寻常的。它由一个由四个同源模块组成的单一多肽组成，每个模块有300个氨基酸残基。大多数对其功能的研究都集中在11-顺式视黄醇和全反式视黄醇在视杆和色素上皮之间的视觉周期运输，以及脂肪酸的运输。 含有全长哺乳动物蛋白质的四个模块首次在脂肪酸配体的存在下结晶。在室内旋转阳极X射线发生器中，晶体在室温下表现出衍射，但在低温冷却时出现了混乱。在试验了多种冷冻降温剂后，在含有20%甘油的情况下，聚乙二醇35K似乎提供了最大的低温保护。我们现在已经从18%的35K聚乙二醇中生长出了晶体。除了常规的数据收集，我们还建议试验由Sol Gruner博士在国际象棋比赛中开发的高压冷冻冷却方法。Gruner博士的团队已经同意在这些实验中协助我们。

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. The 143 kDa human/mammalian interphotoreceptor retinoid-binding protein (IRBP) is unusual in that it has no homology with other retinoid-binding proteins. It consists of a single polypeptide composed of four homologous modules each ~300 amino acid residues. Most studies into its function have focused on the visual cycle trafficking of 11-cis retinal and all-trans retinol between the rods and pigment epithelium, as well as of fatty acids. The four module containing full-length mammalian protein has been crystallized for the first time, in the presence of a fatty acid ligand. The crystals show diffraction at room temperature at the in-house rotating anode X-ray generator, but disorders on cryo-cooling. After experimenting with a number of cryo-cooling agents, PEG 35K seems to offer most cryo-protection, in the presence of 20% glycerol. We have now grown the crystals from 18% 35K PEG. In addition to routine data collection, we propose also to experiment with the high pressure cryo-cooling method developed by Dr. Sol Gruner at CHESS. Dr. Gruner's group has agreed to assist us in these experiments.