Fig. 1.
Glutathionylation of PTP1B is induced by the addition of exogenous H2O2. NR8383 cells were exposed to 100 μM H2O2 and then lysed with a buffer containing N-ethylmaleimide (NEM). GSH was added to remove excess NEM. Whole cell extracts were immunoprecipitated with an antibody to PTP1B. Membranes were immunoblotted with antibodies to either glutathione-protein complexes or PTP1B. Based on the densitometry of IP from two experiments, the maximum percentage PTP1B-SSG in respect to total PTP1B was 31.3 ± 19.1%, which was at 10 min. The results followed the same pattern of increase to a maximum for both experiments. Despite the variability, which is largely due to the semiquantitative nature of densitometry, the pattern of change over time in both experiments was the same.