title: Biosynthesis of intestinal microvillar proteins. Processing of aminopeptidase N by microsomal membranes.
abstract: The biosynthesis of small-intestinal aminopeptidase N (EC 3.4.11.2) was studied in a cell-free translation system derived from rabbit reticulocytes. When dog pancreatic microsomal fractions were present during translation, most of the aminopeptidase N synthesized was found in a membrane-bound rather than a soluble form, indicating that synthesis of the enzyme takes place on ribosomes attached to the rough endoplasmic reticulum. The microsomal fractions process the Mr-115 000 polypeptide, which is the primary translation product of aminopeptidase N, to a polypeptide of Mr 140 000. This was found to be sensitive to the action of endo-beta-N-acetylglucosaminidase H (EC 3.2.1.96), showing that aminopeptidase N undergoes transmembrane glycosylation during synthesis. The position of the signal sequence in aminopeptidase N was determined by a synchronized translation experiment. It was found that microsomal fractions should be added before about 25% of the polypeptide was synthesized to ensure processing to the high-mannose glycosylated form. This suggests that the signal sequence is situated in the N-terminal part of the aminopeptidase N. The size of the cell-free translation product in the absence of microsomal fractions was found to be similar to that on one of the forms of the enzyme obtained from tunicamycin-treated organ-cultured intestinal explants.
mesh_terms: D000118:Acetylglucosaminidase / Q000494:pharmacology; D000626:Aminopeptidases / Q000096:biosynthesis*; D000818:Animals; D018826:CD13 Antigens; D002474:Cell-Free System; D004285:Dogs; D007421:Intestine, Small / Q000378:metabolism*; D007425:Intracellular Membranes / Q000201:enzymology; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D008565:Membrane Proteins / Q000096:biosynthesis*; D008861:Microsomes / Q000201:enzymology; D008871:Microvilli / Q000378:metabolism; D009924:Organ Culture Techniques; D010455:Peptides / Q000378:metabolism; D014176:Protein Biosynthesis; D013552:Swine
chemical_list: D008565:Membrane Proteins; D010455:Peptides; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D000626:Aminopeptidases; D018826:CD13 Antigens