Epinephrine‐Binding Plasma‐Membrane Antigens in Rat Liver

Detergent extracts of isolated rat liver plasma membranes were analysed in two‐dimensional immunoelectrophoresis against antiserum to plasma membranes. Enzyme staining of the immuno‐precipitates revealed the presence of about ten antigens with nucleoside di‐ and triphosphatase activity. Most of these were earlier shown also to be NADH‐neotetrazolium reductase active. In addition, two of these antigens exhibited l ‐leucyl‐β‐naphthylamidase activity. As judged from autoradiography these plasma membrane antigens earlier characterized as multienzyme complexes bound [ 14 C]epinephrine, and the same antigens were labelled regardless of whether membranes or membrane extracts were incubated with the radioactive hormone. The specificity of this binding was established in displacement experiments with unlabelled hormones or their analogues. Another hormone‐binding antigen, also identified in the plasma membrane extract did not exhibit any known enzyme activity while three antigens with different enzyme activities had no epinephrine‐binding capacity. [ 14 C]Epinephrine‐labelled plasma membrane extracts were chromatographed on Sepharose 4B and the fractions obtained were analysed in two‐dimensional immunoelectrophoresis combined with autoradiography. Nucleoside di‐ and triphosphatases of high molecular weights (5) were associated with l ‐leucyl‐β‐naphthylamidase activity, while no such associations were detected in a lower molecular weight region (70000). Further immunological studies on the various fractionated antigens provided evidence that at least two of them occurred in both low and high molecular weight fractions. Hormone‐binding membrane components in varying concentrations were found throughout the eluted extract.