Proteins of the kidney microvillar membrane. Immunoelectrophoretic analysis of the membrane hydrolases: identification and resolution of the detergent- and proteinase-solubilized forms.
Antibodies raised in rabbits to detergent-solubilized pig kidney microvillar proteins have been used to investigate the membrane hydrolases by crossed immunoelectrophoresis. Eight enzymes were detected by specific staining methods: aminopeptidase M, dipeptidylpeptidase IV, neutral endopeptidase, aminopeptidase A, carboxypeptidase P, gamma-glutamyltransferase, trehalase and phosphodiesterase I. The mobility of all these enzymes, with the exception of trehalase and neutral endopeptidase, was increased by treatment of the detergent-solubilized preparation with papain. The difference between the detergent and proteinase forms of these enzymes is attributed to the removal of a small, non-antigenic peptide to which detergent is bound in significant quantities. This interpretation was further supported by experiments in which the microvillus fraction was labelled with an intramembrane photolabelling reagent, 1-azido-4-[125I]iodobenzene. After photolysis, the radioactivity in the membrane could be solubilized by detergent treatment but not by papain treatment. Radioautography after crossed charge-shift immunoelectrophoresis showed a good correlation between charge-shift (signifying the presence of detergent bound to a hydrophobic domain) and the presence of the label.
D000818:Animals; D001386:Azides; D002462:Cell Membrane / Q000201:enzymology*; D055598:Chemical Phenomena; D002621:Chemistry; D003902:Detergents; D006867:Hydrolases / Q000302:isolation & purification*; D007123:Immunoelectrophoresis, Two-Dimensional; D007202:Indicators and Reagents; D007462:Iodobenzenes; D007668:Kidney / Q000201:enzymology*; D008565:Membrane Proteins / Q000032:analysis; D008566:Membranes / Q000201:enzymology; D008871:Microvilli / Q000201:enzymology*; D010206:Papain; D010447:Peptide Hydrolases; D010782:Photolysis; D013552:Swine
D001386:Azides; D003902:Detergents; D007202:Indicators and Reagents; D007462:Iodobenzenes; D008565:Membrane Proteins; D006867:Hydrolases; D010447:Peptide Hydrolases; D010206:Papain