Since the discovery that the endothelium derived relaxing factor (EDRF) was the endogenous toxic gas nitric oxide (NO), an astonishing number of physiological functions have been attributed to NO. Despite the widely recognized importance of NO, little is known about the mechanism of regulation of the NO receptor, the soluble guanylyl cyclase (sGC). sGC is a heme containing heterodimer that catalyzes the formation of cGMP from the substrate GTP. Upon binding of NO, the sGC is activated several hundred fold. The sGC is a multi-domain signaling enzyme that contains the NO receptor-heme domain, a dimerization domain and the effector-catalytic domain. It is not known how the NO signal is propagated to the catalytic domain of sGC. We recently discovered that sGC is desensitized by S-nitrosylation, the addition of a NO moiety to the free thiol of a specific cysteine (Cys). There s now substantial evidence that thiol redox reactions are critical and dynamic regulators of sGC function and that thiol modifications of sGC have clinical relevance as they are associated with decreased vascular reactivity in hypertension. However, the mechanisms by which modification of reactive Cys modulate sGC activation, NO signal transduction to the catalytic domain, sGC basal activity, domain interactions and NO-heme affinity have yet to be explored. The proposed studies seek to understand the structural and molecular basis of mechanisms of regulation of the sGC and how disruption of these mechanisms contributes to development of cardiovascular pathologies. We will use purified enzymes, cellular system and animals models to conduct structure-function studies, molecular dynamics simulation, biochemical and kinetics analysis and applied physiology for the following aims: 1) define the role of thiol Cys in the molecular mechanism of activation of sGC;2) investigate the modulation of sGC by the thiol-reducing protein thioredoxin and 3) establish how thiol-dependent dysfunction of sGC contributes to hypertension and cardiac hypertrophy. Understanding the mechanisms of regulation of sGC will be key to uncovering the molecular basis of some types of hypertension, atherosclerosis and erectile dysfunction, which affect more than 60 millions Americans.

Public Health Relevance

Nitric oxide (NO) induces in blood vessels the production of a small molecule, cGMP, which vasodilates the vasculature. Dysfunction in the NO-cGMP pathway is responsible for many cardiovascular diseases including hypertension, erectile dysfunction and atherosclerosis which affect more than 60 million Americans. We seek to understand how the production of these molecules is controlled by the body.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM067640-11
Application #
8452113
Study Section
Special Emphasis Panel (ZRG1-CB-D (02))
Program Officer
Dunsmore, Sarah
Project Start
2003-04-15
Project End
2013-06-30
Budget Start
2013-04-01
Budget End
2013-06-30
Support Year
11
Fiscal Year
2013
Total Cost
$82,806
Indirect Cost
$29,725
Name
University of Medicine & Dentistry of NJ
Department
Pharmacology
Type
Schools of Medicine
DUNS #
623946217
City
Newark
State
NJ
Country
United States
Zip Code
07107
Heckler, Erin J; Crassous, Pierre-Antoine; Baskaran, Padmamalini et al. (2013) Protein disulfide-isomerase interacts with soluble guanylyl cyclase via a redox-based mechanism and modulates its activity. Biochem J 452:161-9
Duran, Walter N; Beuve, Annie V; Sanchez, Fabiola A (2013) Nitric oxide, S-nitrosation, and endothelial permeability. IUBMB Life 65:819-26
Ramachandran, Jayalakshmi; Schneider, Joel S; Crassous, Pierre-Antoine et al. (2013) Nitric oxide signalling pathway in Duchenne muscular dystrophy mice: up-regulation of L-arginine transporters. Biochem J 449:133-42
Fioramonti, Xavier; Deak, Adam; Deshpande, Srinidhi et al. (2013) Hypothalamic S-nitrosylation contributes to the counter-regulatory response impairment following recurrent hypoglycemia. PLoS One 8:e68709
Baskaran, Padmamalini; Heckler, Erin J; van den Akker, Focco et al. (2011) Aspartate 102 in the heme domain of soluble guanylyl cyclase has a key role in NO activation. Biochemistry 50:4291-7
Wu, Changgong; Parrott, Andrew M; Fu, Cexiong et al. (2011) Thioredoxin 1-mediated post-translational modifications: reduction, transnitrosylation, denitrosylation, and related proteomics methodologies. Antioxid Redox Signal 15:2565-604
Baskaran, Padmamalini; Heckler, Erin J; van den Akker, Focco et al. (2011) Identification of residues in the heme domain of soluble guanylyl cyclase that are important for basal and stimulated catalytic activity. PLoS One 6:e26976
Ma, Xiaolei; Beuve, Annie; van den Akker, Focco (2010) Crystal structure of the signaling helix coiled-coil domain of the beta1 subunit of the soluble guanylyl cyclase. BMC Struct Biol 10:2
Martin, Faye; Baskaran, Padmamalini; Ma, Xiaolei et al. (2010) Structure of cinaciguat (BAY 58-2667) bound to Nostoc H-NOX domain reveals insights into heme-mimetic activation of the soluble guanylyl cyclase. J Biol Chem 285:22651-7
Zhou, Zongmin; Sayed, Nazish; Pyriochou, Anastasia et al. (2008) Protein kinase G phosphorylates soluble guanylyl cyclase on serine 64 and inhibits its activity. Arterioscler Thromb Vasc Biol 28:1803-10

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