Abstract
Sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) is a transmembrane protein that transfers calcium ions from the cytosol of the cell to the lumen of the sarco/endoplasmic reticulum (SR) at the expense of ATP hydrolysis during muscle relaxation. Here we report the electrochemical monitoring of a SERCA1 isoform isolated from rabbit skeletal muscle. The constant-current chronopotentiometric stripping (CPS) analysis was applied for the analysis of solubilized SERCA1 based on a catalytic hydrogen evolution reaction (CHER). Consequently we applied an optimized CPS electrochemical protocol for monitoring the interaction of SERCA1 with the dicarbonyl compounds methylglyoxal, glyoxal and 3‑deoxyglucosone. We observed that SERCA1 readily interacts with methylglyoxal at pH 7.4. Electrochemical analysis showed that 1 mg/ml (9.1 μmol/l) SERCA1 was fully modified by methylglyoxal (3 mmol/l) after 24 h. The dicarbonyl binding decreased the enzyme activity of purified SERCA1 in the following order: methylglyoxal ≫ glyoxal > 3‑deoxyglucosone. The methodology presented here could be used in further studies of the structural integrity and intermolecular interactions of membrane transporters and the study of their oxidative modifications, carbonylation or glycoxidation.
Published Version
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