3-nitrotyrosine and methionine-sulfoxide

Reactive oxygen and nitrogen species can modify various biomolecules, including proteins. The resulting protein modifications are highly diverse, can be reversible as well as irreversible, and might affect protein structure and function. Besides random modifications, targeted modifications at specific amino acids in surface-accessible protein regions occur. These changes are of particular interest as, e.g. by altering the local protein conformation; they might initiate specific (oxidative) signaling pathways. Here, we focus on two protein modifications that are found under conditions of oxidative stress in plants: oxidation of the sulfur-containing methionine and nitration of tyrosine. We review the functional consequences caused by the oxidation of several plant proteins and line-up those proteomics technologies that are amenable to study these selected modifications.

Topics: Animals; Humans; Methionine; Oxidation-Reduction; Plant Proteins; Protein Processing, Post-Translational; Proteomics; Reactive Nitrogen Species; Reactive Oxygen Species; Tandem Mass Spectrometry; Tyrosine

An enhanced formation of reactive oxygen species and peroxynitrite occurs in several clinical settings including diabetes, coronary artery disease, stroke, sepsis, and chronic inflammatory diseases. Peroxynitrite oxidizes methionine and tyrosine residues to methionine sulfoxide (MetSO) and 3-nitrotyrosine (NT), respectively. Notably, ADAMTS-13 cleaves von Willebrand factor (VWF) exclusively at the Tyr1605-Met1606 peptide bond in the A2 domain. We hypothesized that peroxynitrite could oxidize either or both of these amino acid residues, thus potentially affecting ADAMTS-13-mediated cleavage. We tested our hypothesis using synthetic peptide substrates based on: (1) VWF Asp1596-Ala1669 sequence (VWF74) and (2) VWF Asp1596-Ala1669 sequence containing nitrotyrosine (VWF74-NT) or methionine sulfoxide (VWF74-MetSO) at position 1605 or 1606, respectively. The peptides were treated with recombinant ADAMTS-13 and the cleavage products analyzed by RP-HPLC. VWF74 oxidized by peroxynitrite underwent a severe impairment of its hydrolysis. Likewise, VWF74-MetSO was minimally hydrolyzed, whereas VWF74-NT was hydrolyzed slightly more efficiently than VWF74. Oxidation by peroxynitrite of purified VWF multimers inhibited ADAMTS-13 hydrolysis, but did not alter their electrophoretic pattern nor their ability to induce platelet agglutination by ristocetin. Moreover, VWF purified from type 2 diabetic patients showed oxidative damage, as revealed by enhanced carbonyl, NT, and MetSO content and was partially resistant to ADAMTS-13 hydrolysis. In conclusion, peroxynitrite may contribute to prothrombotic effects, hindering the proteolytic processing by ADAMTS-13 of high-molecular-weight VWF multimers, which have the highest ability to bind and activate platelets in the microcirculation.

Topics: ADAM Proteins; ADAMTS13 Protein; Adult; Binding Sites; Blood Platelets; Case-Control Studies; Diabetes Mellitus, Type 2; Female; Humans; Hydrolysis; Male; Methionine; Middle Aged; Oxidative Stress; Peptide Fragments; Peroxynitrous Acid; Platelet Aggregation; Protein Multimerization; Ristocetin; Tyrosine; von Willebrand Diseases; von Willebrand Factor