Cysteine S-nitrosylation protects protein-tyrosine phosphatase 1B against oxidation-induced permanent inactivation.
Protein S-nitrosylation mediated by cellular nitric oxide (NO) plays a primary role in executing biological functions in cGMP-independent NO signaling. Although S-nitrosylation appears similar to Cys oxidation induced by reactive oxygen species, the molecular mechanism and biological consequence remain unclear. We investigated the structural process of S-nitrosylation of protein-tyrosine phosphatase 1B (PTP1B). We treated PTP1B with various NO donors, including S-nitrosothiol reagents and compound-releasing NO radicals, to produce site-specific Cys S-nitrosylation identified using advanced mass spectrometry (MS) techniques. Quantitative MS showed that the active site Cys-215 was the primary residue susceptible to S-nitrosylation. The crystal structure of NO donor-reacted PTP1B at 2.6 A resolution revealed that the S-NO state at Cys-215 had no discernible irreversibly oxidized forms, whereas other Cys residues remained in their free thiol states. We further demonstrated that S-nitrosylation of the Cys-215 residue protected PTP1B from subsequent H(2)O(2)-induced irreversible oxidation. Increasing the level of cellular NO by pretreating cells with an NO donor or by activating ectopically expressed NO synthase inhibited reactive oxygen species-induced irreversible oxidation of endogenous PTP1B. These findings suggest that S-nitrosylation might prevent PTPs from permanent inactivation caused by oxidative stress.
Institute of Biological Chemistry, National Core Facility for Proteomics Research, Academia Sinica, Taipei 11529, Taiwan.
Yi-Yun
YY
Chen
Hsing-Mao
HM
Chu
Kuan-Ting
KT
Pan
Chun-Hung
CH
Teng
Danny-Ling
DL
Wang
Andrew H-J
AH
Wang
Kay-Hooi
KH
Khoo
Tzu-Ching
TC
Meng
D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002522:Chlorocebus aethiops; D018360:Crystallography, X-Ray / Q000379:methods; D003545:Cysteine / Q000737:chemistry*; D004789:Enzyme Activation; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D009569:Nitric Oxide / Q000737:chemistry; D009584:Nitrogen / Q000737:chemistry; D018384:Oxidative Stress; D010100:Oxygen / Q000737:chemistry*; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1 / Q000737:chemistry*; D017382:Reactive Oxygen Species
D017382:Reactive Oxygen Species; D009569:Nitric Oxide; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D003545:Cysteine; D009584:Nitrogen; D010100:Oxygen