The erythrocyte anion transport protein is contranslationally inserted into microsomes.
The biosynthesis of the erythrocyte anion transport protein, Band III (molecular weight 100,000), is of interest, as its NH2-terminal half is hydrophilic and faces the cytoplasmic surface, and its COOH-terminal half spans the phospholipid bilayer several times. To investigate the problem of insertion of Band III into membranes, we used erythroid precursor cells from the spleens of anemic mice as a source of messenger RNA for in vitro studies in the wheat germ and reticulocyte lysate cell-free system containing dog pancreatic microsomes. Immediately after synthesis, Band III was found to be inserted into intracellular membranes in its mature configuration, with the NH2-terminal portion exposed to the cytoplasm and its hydrophobic COOH-terminal portion spanning the lipid bilayer. The newly synthesized Band III was provided with a high mannose asparagine-linked oligosaccharide, which was sensitive to cleavage by endoglycosidase H; this is presumably the precursor of the very large and complex oligosaccharide found on the finished molecule. Band III was found to insert into dog pancreatic microsomes in a cotranslational manner; in synchronized translation studies microsomes could be added as late as the time when the hydrophilic NH2-terminal half of the protein had been synthesized and still allow normal transmembrane insertion and glycosylation. There is no cleavage of any NH2-terminal peptide during membrane insertion. The results suggest that Band III contains a sequence near the middle of the protein that directs its insertions into endoplasmic reticulum membranes.
D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte; D001798:Blood Proteins / Q000096:biosynthesis*; D002474:Cell-Free System; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D006023:Glycoproteins / Q000096:biosynthesis*; D008565:Membrane Proteins / Q000096:biosynthesis*; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000378:metabolism
D001457:Anion Exchange Protein 1, Erythrocyte; D001798:Blood Proteins; D006023:Glycoproteins; D008565:Membrane Proteins; D011498:Protein Precursors