TY - JOUR
T1 - Superoxide-induced stimulation of protein kinase C via thiol modification and modulation of zinc content
AU - Knapp, Lauren T.
AU - Klann, Eric
PY - 2000/8/4
Y1 - 2000/8/4
N2 - We investigated the effects of mild oxidation on protein kinase C (PKC) using the xanthine/xanthine oxidase system of generating superoxide. Exposure of various PKC preparations to superoxide stimulated the autonomous activity of PKC. Similarly, thiol oxidation increased autonomous PKC activity, consistent with the notion that superoxide stimulates PKC via thiol oxidation. The superoxide-induced stimulation of PKC activity was partially reversed by reducing agents, suggesting that disulfide bond formation contributed to the oxidative stimulation of PKC. In addition, superoxide increased the autonomous activity of the α, β(II), ε, and ζ PKC isoforms, all of which contain at least one cysteinerich region. Taken together, our observations suggested that superoxide interacts with PKC at the cysteine-rich region, zinc finger motif of the enzyme. Therefore, we examined the effects of superoxide on this region by testing the hypothesis that superoxide stimulates PKC by promoting the release of zinc from PKC. We found that a zinc chelator stimulated the autonomous activity of PKC and that superoxide induced zinc release from an PKC-enriched enzyme preparation. In addition, oxidized PKC contained significantly less zinc than reduced PKC. Finally, we have isolated a persistent, autonomously active PKC by DEAE-cellulose column chromatography from hippocampal slices incubated with superoxide. Taken together, these data suggest that superoxide stimulates autonomous PKC activity via thiol oxidation and release of zinc from cysteine-rich region of PKC.
AB - We investigated the effects of mild oxidation on protein kinase C (PKC) using the xanthine/xanthine oxidase system of generating superoxide. Exposure of various PKC preparations to superoxide stimulated the autonomous activity of PKC. Similarly, thiol oxidation increased autonomous PKC activity, consistent with the notion that superoxide stimulates PKC via thiol oxidation. The superoxide-induced stimulation of PKC activity was partially reversed by reducing agents, suggesting that disulfide bond formation contributed to the oxidative stimulation of PKC. In addition, superoxide increased the autonomous activity of the α, β(II), ε, and ζ PKC isoforms, all of which contain at least one cysteinerich region. Taken together, our observations suggested that superoxide interacts with PKC at the cysteine-rich region, zinc finger motif of the enzyme. Therefore, we examined the effects of superoxide on this region by testing the hypothesis that superoxide stimulates PKC by promoting the release of zinc from PKC. We found that a zinc chelator stimulated the autonomous activity of PKC and that superoxide induced zinc release from an PKC-enriched enzyme preparation. In addition, oxidized PKC contained significantly less zinc than reduced PKC. Finally, we have isolated a persistent, autonomously active PKC by DEAE-cellulose column chromatography from hippocampal slices incubated with superoxide. Taken together, these data suggest that superoxide stimulates autonomous PKC activity via thiol oxidation and release of zinc from cysteine-rich region of PKC.
UR - http://www.scopus.com/inward/record.url?scp=0034604723&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0034604723&partnerID=8YFLogxK
U2 - 10.1074/jbc.M002043200
DO - 10.1074/jbc.M002043200
M3 - Article
C2 - 10823825
AN - SCOPUS:0034604723
SN - 0021-9258
VL - 275
SP - 24136
EP - 24145
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 31
ER -