Synthesis of α1‐Antitrypsin in Rat‐Liver Hepatocytes and in a Cell‐Free System

The biosynthesis of α1‐antitrypsin, a secretory glycoprotein, has been studied in vitro in a cell‐free translation system from wheat germ and in vivo with hepatocytes. When messenger RNA from rat liver polysomes was translated in a heterologous cell‐free system a translation product was immunoprecipitated by anti‐(α1‐antitrypsin) which had a smaller molecular weight (42000) on sodium dodecyl sulfate/polyacrylamide gel electrophoresis than the authentic glycoprotein (54000) isolated from rat serum. An intact amino terminus of the α1‐antitrypsin synthesized in vitro could be demonstrated by repetitive Edman degradation.When hepatocytes, labeled with [35S]methionine, were analyzed with respect to newly synthesized α1‐antitrypsin, a protein with a molecular weight of 49000 was immunoprecipitated from a high‐speed supernatant. When the hepatocytes were treated with tunicamycin, an inhibitor of protein glycosylation, a protein of a molecular weight of 40000 was immunoprecipitated from the cells as well as from the medium. Interestingly, an α1‐antitrypsin form of an even higher molecular weight (54000) was immunoprecipitated from the hepatocyte monolayer medium. Several conclusions can be drawn from these findings. First, α1‐antitrypsin is synthesized as a larger‐molecular‐weight precursor with an extra amino‐terminal sequence of about 20 amino acids. Second, the liver form of α1‐antitrypsin is different from the α1‐antitrypsin which has been secreted, and third, the carbohydrate moieties of rat liver α1‐antitrypsin do not appear to represent an absolute requirement for the secretion process.A length of 1930 ± 170 nucleotides was estimated for the poly(A)‐rich mRNA of the precursor of rat liver α1‐antitrypsin; a non‐coding region of about 700 nucleotides must exist.Furthermore, it is shown that the major part of the poly(A)‐rich RNA for α1‐antitrypsin is found in loosely and tightly membrane‐bound polyribosomes. This finding is in agreement with the current ideas on the site of synthesis of secretory proteins.