title: Identification of novel S-nitrosation sites in soluble guanylyl cyclase, the nitric oxide receptor.
abstract: Soluble Guanylyl Cyclase (sGC) is the main receptor for nitric oxide (NO). NO activates sGC to synthesize cGMP, triggering a plethora of signals. Recently, we discovered that NO covalently modifies select sGC cysteines via a post-translational modification termed S-nitrosation or S-nitrosylation. Earlier characterization was conducted on a purified sGC treated with S-nitrosoglutathione, and identified three S-nitrosated cysteines (SNO-Cys). Here we describe a more biologically relevant mapping of sGC SNO-Cys in cells to better understand the multi-faceted interactions between SNO and sGC. Since SNO-Cys are labile during LC/MS/MS, MS analysis of nitrosation typically occurs after a biotin switch reaction, in which a SNO-Cys is converted to a biotin-Cys. Here we report the identification of ten sGC SNO-Cys in rat neonatal cardiomyocytes using an Orbitrap MS. A majority of the SNO-Cys identified is located at the solvent-exposed surface of the sGC, and half of them in the conserved catalytic domain, suggesting biological significance. These findings provide a solid basis for future studies of the regulations and functions of diverse sGC S-nitrosation events in cells.
authors.affiliation: Department of Pharmacology, Physiology and Neuroscience, Rutgers University, New Jersey Medical School, Newark, NJ 07103, United States.
authors.forename: Annie
authors.initials: A
authors.lastname: Beuve
authors.affiliation: Center for Advanced Proteomics Research, Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers University, New Jersey Medical School Cancer Center, Newark, NJ 07103, United States.
authors.forename: Changgong
authors.initials: C
authors.lastname: Wu
authors.affiliation: Center for Advanced Proteomics Research, Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers University, New Jersey Medical School Cancer Center, Newark, NJ 07103, United States.
authors.forename: Chuanlong
authors.initials: C
authors.lastname: Cui
authors.affiliation: Center for Advanced Proteomics Research, Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers University, New Jersey Medical School Cancer Center, Newark, NJ 07103, United States.
authors.forename: Tong
authors.initials: T
authors.lastname: Liu
authors.affiliation: Center for Advanced Proteomics Research, Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers University, New Jersey Medical School Cancer Center, Newark, NJ 07103, United States.
authors.forename: Mohit Raja
authors.initials: MR
authors.lastname: Jain
authors.affiliation: Department of Pharmacology, Physiology and Neuroscience, Rutgers University, New Jersey Medical School, Newark, NJ 07103, United States.
authors.forename: Can
authors.initials: C
authors.lastname: Huang
authors.affiliation: Center for Advanced Proteomics Research, Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers University, New Jersey Medical School Cancer Center, Newark, NJ 07103, United States.
authors.forename: Lin
authors.initials: L
authors.lastname: Yan
authors.affiliation: High Performance and Research Computing, OIRT, Rutgers University, New Brunswick, NJ 07103, United States; Department of Pharmacology, Rutgers University, Robert Wood Johnson Medical School, Piscataway, NJ.
authors.forename: Vladyslav
authors.initials: V
authors.lastname: Kholodovych
authors.affiliation: Center for Advanced Proteomics Research, Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers University, New Jersey Medical School Cancer Center, Newark, NJ 07103, United States. Electronic address: liho2@rutgers.edu.
authors.forename: Hong
authors.initials: H
authors.lastname: Li
mesh_terms: D000818:Animals; D003545:Cysteine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000201:enzymology*; D009569:Nitric Oxide / Q000737:chemistry / Q000378:metabolism*; D015538:Nitrosation; D051381:Rats; D017208:Rats, Wistar; D026422:S-Nitrosoglutathione / Q000737:chemistry / Q000378:metabolism; D026403:S-Nitrosothiols / Q000737:chemistry / Q000378:metabolism; D000071756:Soluble Guanylyl Cyclase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D026403:S-Nitrosothiols; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione; C021315:S-nitrosocysteine; D000071756:Soluble Guanylyl Cyclase; D003545:Cysteine
keywords: Biotin switch; S-nitrosation; Soluble guanylyl cyclase; Tandem mass spectrometry