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pmid:25181601
Differential cysteine labeling and global label-free proteomics reveals an altered metabolic state in skeletal muscle aging.
Brian McDonagh · Giorgos K Sakellariou · Neil T Smith · Philip Brownridge · Malcolm J Jackson
Journal of proteome research · 2014 doi:10.1021/pr5006394
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Complete file
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Reactive oxygen and reactive nitrogen species (ROS/RNS) generated by contracting skeletal muscle are necessary for optimal protein function, signaling, and adaptation. To investigate the redox proteome of aging gastrocnemius muscles from adult and old male mice, we developed a label-free quantitative proteomic approach that includes a differential cysteine labeling step. The approach allows simultaneous identification of up- and downregulated proteins between samples in addition to the identification and relative quantification of the reversible oxidation state of susceptible redox cysteine residues. Results from muscles of adult and old mice indicate significant changes in the content of chaperone, glucose metabolism, and cytoskeletal regulatory proteins, including Protein DJ-1, cAMP-dependent protein kinase type II, 78 kDa glucose regulated protein, and a reduction in the number of redox-responsive proteins identified in muscle of old mice. 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Park7 has previously been reported in vivo and in vitro to be a target of S-nitrosylati", "dissected. A gastrocnemius muscle from each mouse was placed immediately in thiol blocking buf", "corresponding gastrocnemius muscle from each mouse was snap-frozen in liquid nitrogen and store"], "order_basis": "paper_text_order:1791", "id": "assay:1"}, {"role": "data_generation", "definition_id": "western_blot", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["y 0.8, and at least one unique peptide). (B) Western blotting for Grp78, Gst P1, Hsp70, and Cryab. (C) Str", "ocnemius muscle from adult and old mice. (B) Western blotting validation of aconitase expression. (C) Acon", "date the label-free quantification approach, immunoblotting of a number of the differentially regulated", "ated proteins was performed. 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