Roles of superoxide radical anion in signal transduction mediated by reversible regulation of protein-tyrosine phosphatase 1B

Growth factors induce intracellular production of reactive oxygen species i n non-phagocytic cells and elevation of their phosphorylated protein tyrosi ne level. The latter can be achieved by activating protein-tyrosine kinases and/or inactivating protein-tyrosine phosphatases (PTPs). A highly abundan t PTP, PTP-1B, is known to be inactivated by oxidation of its catalytic sit e Cys-215. We show that O-2 is kinetically more efficient and chemically mo re specific oxidant than H2O2 for inactivating PTP-1B. The second-order rat e constant for the O-2- and H2O2-mediated inactivation is 334 +/- 45 M-1 s( -1) and 42.8 +/- 3.8 M-1 s(-1), respectively. PTP-1B oxidized by H2O2 exhib its significantly more oxidized methionine residues and shows a lower degre e of reversibility. The initial oxidative product, the Cys-215 sulfenic der ivative, can easily be oxidized further to its irreversible sulfinic and su lfonic derivatives. This step is prevented by glutathionylation of the sulf enic derivative to form a S-glutathionylated PTP-1B, which can be reactivat ed by dithiothreitol or thioltransferase. Thus, a signal transduction mecha nism mediated by the O-2(radical anion) and the participation of glutathion e is proposed for the regulation of PTP-1B. This mechanism is supported by the in vivo demonstration that glutathionylated PTP-1B at Cys-215 is formed in A431 cells when they were treated with epidermal growth factor.