Identification of the protein responsible for hepatic system N amino acid transport activity.
In the liver, glutamine utilization may be limited by the rate of transport across the plasma membrane by the System N carrier. System N-mediated transport activity has been solubilized from rat liver plasma membrane, partially purified, and then reconstituted into proteoliposomes. To identify the System N carrier protein, monoclonal antibodies were generated against the protein fraction enriched for System N activity. Two antibodies , 3E1-2 and 1E7-3, inhibited System N activity in hepatocytes. These antibodies also immunoprecipitated System N activity from a mixture of solubilized proteins and were specific for antigen recognition in that neither immunoprecipitated System A activity. The antibody recognized a single protein of molecular size 100 kDa by immunoblot analysis. Recognition of this protein by the antibody increased in parallel with the enrichment of System N activity in solubilized membrane fractions. These data suggest that a 100-kDa plasma membrane protein mediates System N transport activity in rat hepatocytes.
Tamarappoo
B K
BK
Department of Biochemistry and Molecular Biology, J. Hillis Miller Health Center, University of Florida, Gainesville 32610.
Handlogten
M E
ME
Laine
R O
RO
Serrano
M A
MA
Dugan
J
J
Kilberg
M S
MS
D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D001692:Biological Transport; D015153:Blotting, Western; D002352:Carrier Proteins / Q000032:analysis* / Q000276:immunology; D002462:Cell Membrane / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D008099:Liver / Q000737:chemistry* / Q000378:metabolism; D011233:Precipitin Tests; D051381:Rats
D000596:Amino Acids; D000911:Antibodies, Monoclonal; D002352:Carrier Proteins