Purification, characterization and origin of rat gastric peroxidase.
A membrane-bound peroxidase (EC 1.11.1.7) from rat stomach has been solubilized by 0.2% cetyltrimethylammonium bromide in the presence of 1.2 M NH4Cl. The enzyme was purified 3355-fold to apparent homogeneity as judged by acid polyacrylamide gel electrophoresis and appears to be a cationic protein. In sodium dodecyl sulfate gel electrophoresis, the enzyme shows single polypeptide band of Mr 45,000. In gel permeation, the Mr has been estimated as 47,000. Spectral properties indicate the presence of Soret band at 412 nm which shifts to 425 nm on complexation with CN- and to 430 nm on reduction with dithionite. The velocity constant, k1 for the reaction of the peroxidase with H2O2 is 1.38 X 10(7) M-1 s-1 and Km for H2O2 is 0.1 mM. The enzyme contains active sulphydryl groups and is inhibited by sulphydryl reagents of which p-hydroxymercuribenzoate is more reactive than mersalyl or N-ethylmaleimide. The enzyme is very resistant to thermal denaturation up to 65 degrees C and also to chaotropic reagents at least up to 2 M above which it is inactivated. The enzyme shows similarity with the intestinal eosinophil peroxidase as regards the molecular mass, spectral, kinetic and some of the catalytic properties. However, they differ significantly in terms of their interaction with fluoride ion, sulphydryl reagents, chaotropic reagent and also with the antiserum against the gastric peroxidase. Histochemically, the gastric peroxidase is shown to be localised in the gastric gland proper of the fundic stomach, rich in parietal and chief cells.
D000818:Animals; D002384:Catalysis; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005753:Gastric Mucosa / Q000201:enzymology*; D006651:Histocytochemistry; D007158:Immunologic Techniques; D007527:Isoenzymes / Q000302:isolation & purification*; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D009195:Peroxidase; D010544:Peroxidases / Q000302:isolation & purification*; D051381:Rats; D012995:Solubility
D007527:Isoenzymes; D010544:Peroxidases; D009195:Peroxidase