Association of renal dipeptidase with the Triton-insoluble fraction of kidney microvilli.
Renal dipeptidase, previously identified as a component of renal microvilli, has been investigated to determine its orientation within this organelle. Digestion of porcine renal microvilli with papain released essentially all of aminopeptidase M, an outer membrane marker enzyme from the microvilli within one hour; whereas less than 10% of renal dipeptidase was released under the same conditions. Antibody to purified renal dipeptidase produced 50% inhibition of the purified enzyme at an antibody/antigen molar ratio of 2:1. Inhibition by the renal dipeptidase-directed antibody was not observed when the enzyme was bound within the microvillar structure. Demembranation of the microvilli with Triton X-100 resulted in a distribution of 68% of renal dipeptidase in the insoluble pellet and 32% in the soluble supernatant. The same detergent treatment released 92% of animopeptidase M into the supernatant. These results indicate that renal dipeptidase is not located at the luminal surface of the microvillus membrane where it would be available for release of papain, inhibition by antibody, or solubilization by detergent. Fractionation of the Triton-insoluble pellet with 2 M NaCl resulted in the release of 64% of the peptidase into a pellicle fraction separated from insoluble pellet and soluble supernatant. Finally extraction of Triton-insoluble pellet with 0.05 mM ATP-0.10 mM MgCl2 X 6 H2O solubilized 57% of the renal dipeptidase.
D000818:Animals; D004150:Dipeptidases / Q000302:isolation & purification / Q000378:metabolism*; D007672:Kidney Cortex / Q000201:enzymology*; D008871:Microvilli / Q000201:enzymology; D017830:Octoxynol; D011092:Polyethylene Glycols; D012995:Solubility; D013552:Swine / Q000378:metabolism
D011092:Polyethylene Glycols; D017830:Octoxynol; D004150:Dipeptidases; C038937:dipeptidase