title: SNO spectral counting (SNOSC), a label-free proteomic method for quantification of changes in levels of protein S-nitrosation.
abstract: S-Nitrosation plays an important role in regulation of protein function and signal transduction. Discovering S-nitrosated targets is a prerequisite for further functional study. However, current proteomic methods used to quantify S-nitrosation are limited in their applicability to certain types of samples, or by the need for special reagents and complex procedures to obtain the results. Here we devised a label-free proteomic method for quantification of changes in the level of protein S-nitrosation on the basis of a spectral counting strategy, called S-nitrosothiol (SNO) spectral counting (SNOSC). With this method, samples can be from any source (cells, tissues); there is no need for labelling reagents or procedures, and the results yield quantitative information. Moreover, as it is based on the irreversible biotinylation procedure (IBP) for S-nitrosation protein enrichment, false positive targets caused by the interference of intermolecular disulphide bonds are ruled out. Using SNOSC we studied S-nitrosation in the cell line RAW264.7 induced exogenously with S-nitrosoglutathione (GSNO), or induced endogenously by lipopolysaccharides/interferon-gamma (LPS/IFN-γ). We detected a significant increase in S-nitrosation of 50 proteins after exogenous induction and 17 proteins after endogenous induction. We thus demonstrate that SNOSC is a widely applicable proteomic method for fast screening of SNO proteins.
authors.affiliation: National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Chaoyang District, Beijing, China.
authors.forename: Xu
authors.initials: X
authors.lastname: Zhang
authors.forename: Bo
authors.initials: B
authors.lastname: Huang
authors.forename: Chang
authors.initials: C
authors.lastname: Chen
mesh_terms: D000818:Animals; D019921:Biotinylation; D002460:Cell Line; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism; D007371:Interferon-gamma / Q000378:metabolism / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000378:metabolism / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D015233:Models, Statistical; D009569:Nitric Oxide / Q000378:metabolism; D015538:Nitrosation; D011506:Proteins / Q000032:analysis* / Q000378:metabolism; D040901:Proteomics; D026422:S-Nitrosoglutathione / Q000494:pharmacology; D026403:S-Nitrosothiols / Q000378:metabolism
chemical_list: D004220:Disulfides; D008070:Lipopolysaccharides; D011506:Proteins; D026403:S-Nitrosothiols; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione; D007371:Interferon-gamma; D003545:Cysteine