Soluble guanylyl cyclase: mechanisms of activation and modulation

描述（由申请人提供）：可溶性鸟苷酸环化酶（sGC）是一种关键信号蛋白，在被一氧化氮激活时刺激第二信使cGMP的产生。这种cGMP信号传导途径对于许多心血管过程（包括血压调节）是重要的，因此sGC作为治疗心血管疾病（例如高血压、心力衰竭、动脉粥样硬化、勃起功能障碍、血栓形成和肾纤维化）的药物靶标引起了相当大的关注。靶向sGC的几种化合物处于临床前或临床试验阶段。申请人通过确定4个不同结构域的晶体结构，包括与sGC激活剂BAY 58-2667和一氧化氮的复合物，帮助理解鸟苷酸环化酶的结构。本申请的首要目标是进一步揭示sGC的信号传导复杂性，这可以产生对如何利用或绕过该信号传导机制的见解，使得该信息可以在药学上用于调节sGC。拟议的研究是一个高度合作和多学科的性质，涉及技术，如X射线晶体学，电子顺磁共振（EPR），等温量热法（ITC），诱变，和细胞生物学/活性测量。三个具体目标是：具体目标1：为了检验sGC的H-NOX结构域的激活涉及血红素口袋的部分变形的假设。还将探索sGC激活剂的激活以进行比较并描绘这些血红素模拟物的作用模式。具体目标二：检验sGC的卷曲螺旋（CC）结构域是平行二聚体，并且该结构域不是静态的，在活化过程中发生构象变化的假设。具体目标3：为了检验sGC的N-末端一半负责结合化学上不同的sGC刺激物的假设;该sGC区域包括H-NOX和PAS/H-NOXA结构域。拟议的研究将导致对sGC功能和GC激活剂/调节剂如何工作的新分子见解，这将增强我们对心血管过程的理解，并可能导致开发治疗心血管疾病的新药。

DESCRIPTION (provided by applicant): The soluble guanylyl cyclase (sGC) is a key signaling protein that stimulates the production of the second messenger cGMP when activated by nitric oxide. This cGMP signaling pathway is important for a number of cardiovascular processes including blood pressure regulation and sGC has therefore attracted considerable attention as a pharmaceutical drug target to treat cardiovascular diseases such as hypertension, heart failure, atherosclerosis, erectile dysfunction, thrombosis, and renal fibrosis. Several compounds targeting sGC are in pre-clinical or clinical trial stages. The applicant has aided the structural understanding of guanylyl cyclases by determining crystal structures of 4 different domains including complexes with the sGC activators BAY 58-2667 and nitric oxide. The overarching goal of this application is to further unravel the signaling intricacies of sGC that could yield insights into how to either utilize or bypass this signaling mechanism such that this information can be pharmaceutically exploited to modulate sGC. The proposed research is of a highly collaborative and multi-disciplinary nature involving techniques such as X-ray crystallography, electron paramagnetic resonance (EPR), isothermal calorimetry (ITC), mutagenesis, and cell biology/activity measurements. The 3 Specific Aims are: Specific Aim 1: To test the hypothesis that the activation of the H-NOX domain of sGC involves a partial distortion of the heme pocket. The activation by sGC activators will also be probed for comparison and to delineate the mode of action of these heme mimetics. Specific Aim 2: To test the hypothesis that the coiled-coiled (CC) domain of sGC is a parallel dimer and that this domain is not static and undergoes conformational changes during the activation process. Specific Aim 3: To test the hypothesis that the N-terminal half of sGC is responsible for binding sets of chemically distinct sGC stimulators; this sGC region includes the H-NOX and PAS/H-NOXA domain(s). The proposed research will lead to new molecular insights into how sGC functions and how GC activators/modulators work and this will enhance our understanding of cardiovascular processes and could lead to the development of new drugs to treat cardiovascular diseases.