Characterization of the membrane-associated thiol oxidase activity of rat small-intestinal epithelium.

The membrane-associated thiol oxidase of rat small-intestinal epithelium was studied to determine its subcellular localization and properties. The brush-border and basal-lateral regions of the plasma membrane were isolated by density-gradient centrifugation in Percoll. The intestinal oxidase was localized by use of marker enzymes to the basal-lateral region of the plasma membrane. The reaction stoichiometry and activity with a variety of low-molecular-weight thiols were determined. The oxidase activity was inhibited by EDTA, bathocuproine disulfonate, N-ethylmaleimide, and H2O2; this suggests that copper and a sulfhydryl group are involved in catalysis. Oxidase activity in EDTA-treated basal-lateral membranes was reconstituted with CuSO4, which suggests the requirement for copper. These results show that the intestinal oxidase is very similar to the renal oxidase, and because of the subcellular localization and accessibility to extracellular thiols, suggests that the intestinal oxidase may be important in the maintenance of the plasma thiol:disulfide ratio.