Quantification of carbonate radical formation by the bicarbonate-dependent peroxidase activity of superoxide dismutase 1 using pyrogallol red bleaching.

ABSTRACT: Carbonate radicals (CO₃^-) are generated by the bicarbonate-dependent peroxidase activity of cytosolic superoxide dismutase (Cu,Zn-SOD, SOD-1). The present work explored the use of bleaching of pyrogallol red (PGR) dye to quantify the rate of CO₃^- formation from bovine and human SOD-1 (bSOD-1 and hSOD-1, respectively). This approach was compared to previously reported methods using electron paramagnetic resonance spin trapping with DMPO, and the oxidation of ABTS (2,2-azino-bis(3-ethylbenzothiazoline)-6-sulfonic acid). The kinetics of PGR consumption elicited by CO₃^- was followed by visible spectrophotometry. Solutions containing PGR (5-200 μM), SOD-1 (0.3-3 μM), H₂O₂ (2 mM) in bicarbonate buffer (200 mM

SUBMITTER: Figueroa JD 

PROVIDER: S-EPMC6523824 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature