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Cysteinylation of the ALS‐Associated Protein SOD1 Confers Resistance to Oxidation
Author(s) -
Auclair Jared,
Ringe Dagmar,
Petsko Gregory,
Agar Jeffrey
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.717.1
Subject(s) - chemistry , sod1 , superoxide , superoxide dismutase , reactive oxygen species , hydrogen peroxide , biochemistry , peroxide , dimer , dismutase , yeast , cysteine , enzyme , organic chemistry
Natural by‐products of aerobic metabolism are reactive oxygen species (ROS). ROS arise from the incomplete reduction of molecular oxygen, resulting in the production of hydrogen peroxide (H 2 O 2 ) and the superoxide anion (O 2 ‐. ). Cells combat harmful ROS species with a multi‐faceted anti‐oxidant defense mechanism that includes catalytic removal of ROS by enzymes such as copper‐zinc superoxide dismutase (SOD1). Loss of SOD1 function ‐ by oxidation, for example ‐ can cause an increase in superoxide anions, leading to such negative‐effects as neurotoxicity. Preventing oxidation may aid in preventing these negative effects. Recombinant human SOD1 was expressed and purified from yeast and brain or spinal cord either aerobically or anaerobically. Profile spectra of unmodified versus modified SOD1 samples were obtained using a 9.4T FTMS. The 2.5 Å resolution crystal structure of cysteinylated SOD1 (SOD1cys) was determined using molecular replacement. Profile spectra showed post‐translational modifications (PTMs) of SOD1 were present, and that cysteinylation and peroxide‐mediated oxidations appear to account for the majority of observed in vivo PTMs. In addition, we observed that SOD1cys is protected from oxidation in vitro and localized the site of cysteinylation to Cysteine 111. This was further confirmed by the crystal structure, which showed only one cysteine modification per dimer. In addition, we demonstrated SOD1 purified anaerobically had a 50‐fold reduction in cysteinylation compared to an aerobically purified sample, thus suggesting some of the S‐thiolation modifications may be artifacts. We conclude SOD1 cysteinylation is one possible way to combat the negative effects associated with life in an oxidizing environment.