The existence and properties of two dimers of rat liver ecto-5'-nucleotidase.
Immunoaffinity-purified rat liver 5'-nucleotidase contained two subunits of Mr 70 000 (alpha) and 38 000 (beta). Charge-shift electrophoresis and chemical cross-linking revealed that approx. 80% of the solubilized enzyme activity occurred as an alpha alpha-dimer of Mr 140 000. The remaining 20% was an alpha beta-dimer of Mr 108 000. The beta-subunit did not possess enzymic activity. Peptide mapping and immunoblotting with antibodies against the alpha- and beta-subunits showed that the beta-subunit was homologous with a part of the alpha-subunit. Three monoclonal antibodies against rat liver 5'-nucleotidase were characterized as binding to the extracellular domain of the enzyme. All three monoclonal antibodies and concanavalin A bound to the alpha-subunit, but no binding could be detected to the beta-subunit. It was therefore concluded that the beta-subunit was a fragment of an alpha-subunit that had lost an extracellular domain. Both forms of the enzyme occurred in freshly solubilized membrane preparations as well.
D015720:5'-Nucleotidase; D000818:Animals; D000906:Antibodies; D002462:Cell Membrane / Q000201:enzymology; D002846:Chromatography, Affinity; D003208:Concanavalin A; D004120:Dimethyl Suberimidate; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes / Q000276:immunology; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D009708:Nucleotidases / Q000276:immunology; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding; D051381:Rats; D011915:Rats, Inbred BUF
D000906:Antibodies; D007527:Isoenzymes; D046911:Macromolecular Substances; D010446:Peptide Fragments; D003208:Concanavalin A; D004120:Dimethyl Suberimidate; D009708:Nucleotidases; D015720:5'-Nucleotidase