Glutathione reductase in the sea urchin egg. III. Activation of the complex form by proteinases.
The G-200 flow-through fraction of the extract of sea urchin eggs contained a complex form of glutathione reductase (GR) [EC 1.6.4.2]. The complex was unstable and gradually dissociated with ain increase in GR activity. The activation was facilitated by high concentrations of EDTA, KCI or (NH4)2SO4. The rate of activation by salts was apparently dependent on the ionic strength. The complex form was also activated rather quickly by treatment with proteinases such as trypsin [EC 3.4.21.4], alpha-chymotrypsin [EC 3.4.21.1] or subtilisin [EC 3.4.21.14]. Trypsin caused the complex to release the free form of GR. Even after trypsin treatment, little change was observed in the dependence of the GR activity on GSSG or NADPH concentration. The GR activity of the complex form was not inhibited at all by 0.2 mM N-ethylmaleimide (NEM) in the presence of GSSG, but was reduced to 3% in the presence of NADPH. When excess NEM was sequestered with GSH, the NEM-treated complex form was strikingly activated by trypsin, while no activation was detected with the free form of enzyme pretreated with NEM. These results suggest that the active site of GR in the complex form is largely masked by a polypeptide moiety of theinhbitiory component.
D000818:Animals; D001665:Binding Sites; D002918:Chymotrypsin / Q000494:pharmacology; D004789:Enzyme Activation; D005033:Ethylmaleimide / Q000494:pharmacology; D005260:Female; D005978:Glutathione / Q000494:pharmacology; D005980:Glutathione Reductase / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D009249:NADP / Q000494:pharmacology; D010063:Ovum / Q000201:enzymology*; D010084:Oxidation-Reduction; D010447:Peptide Hydrolases / Q000494:pharmacology*; D012617:Sea Urchins; D013381:Subtilisins / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology
D009249:NADP; D005980:Glutathione Reductase; D010447:Peptide Hydrolases; D013381:Subtilisins; D002918:Chymotrypsin; D014357:Trypsin; D005978:Glutathione; D005033:Ethylmaleimide