Protein microarray characterization of the S-nitrosoproteome

Area of Science:

• Proteomics
• Biochemistry
• Molecular Biology

Background:

• Nitric oxide (NO) exerts significant physiological effects through S-nitrosylation.
• The specific cellular targets and modification sites of NO-mediated S-nitrosylation remain largely uncharacterized.

Purpose of the Study:

• To comprehensively identify and characterize the S-nitrosoproteome.
• To define specific cysteine residues modified by S-nitrosylation.
• To uncover novel regulatory pathways influenced by S-nitrosylation.

Main Methods:

• Utilized a high-density protein microarray containing 16,368 unique human proteins.
• Employed a specific labeling and affinity capture technique to identify S-nitrosylated proteins.
• Analyzed protein sequences to identify consensus motifs for S-nitrosylation.

Main Results:

• Identified 834 potentially S-nitrosylated human proteins.
• Pinpointed 138 cysteine residues on 131 peptides within 95 proteins as critical sites of NO action, with 113 being novel.
• Discovered a consensus sequence motif predictive of S-nitrosylation.
• Found that eight ubiquitin E3 ligases are modulated by S-nitrosylation, impacting the ubiquitin-proteasome system.

Conclusions:

• This study defines an extensive landscape of proteins regulated by NO via S-nitrosylation.
• Identified novel sites and a consensus motif for S-nitrosylation, advancing our understanding of NO signaling.
• Revealed S-nitrosylation as a regulatory mechanism for ubiquitin E3 ligases, impacting protein degradation pathways.