STUDIES OF CONFORMATIONAL CHANGES IN SOLUBLE GUANYLATE CYCLASE

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 可溶性鸟苷酸环化酶（sGC）是一种分子量为150 kD的异源二聚体蛋白，是细胞内主要的一氧化氮（NO）受体。NO与β亚基的N-末端结构域中的血红素结合，刺激cGMP的产生并导致NO依赖性信号级联。 体外研究表明，当配体结合时，可能发生变构构象变化。 除了生理配体NO之外，还有效应配体，典型的是YC-1，其与一氧化碳（CO）协同作用。 我们要求光束时间进行小角度X射线散射研究可溶性鸟苷酸环化酶，以表征配体结合相关的构象变化。 人sGC和烟草天蛾sGC的几个不同结构域和构建体已经通过重组方法表达，并且是可用的，或将在不久的将来变得可用。 初步分析超离心研究表明，构象变化足够大，其特征在于由SAXS和蛋白质不聚集在离心实验的条件下显着。

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. Soluble guanylate cyclase (sGC), a 150 kD heterodimeric protein, is the primary nitric oxide (NO) receptor in the cell. NO binds to heme in the N-terminal domain of the beta subunit, stimulates the production of cGMP and leads to NO-dependent signaling cascades. In vitro studies have shown that a probably allosteric conformational change takes place when ligands bind. In addition to the physiological ligand NO, there are effector ligands, typified by YC-1 that act in concert with carbon monoxide (CO). We request beam time to perform small angle x-ray scattering studies on soluble guanylate cyclase in order to characterize ligand binding associated conformational changes. Several different domains and constructs of both human and Manduca sexta sGC have been expressed by recombinant methods and are available, or will become available in the near future. Preliminary analytical ultracentrifugation studies demonstrate that the conformational change is large enough to be characterized by SAXS and that the protein does not aggregate significantly under the conditions of the centrifugation experiment.