Biosynthesis of intestinal microvillar proteins. The effect of swainsonine on post-translational processing of aminopeptidase N

• Published in: Biochemical journal Vol. 216; no. 2; pp. 325 - 331
• Main Authors: Danielsen, E M, Cowell, G M, Norén, O, Sjöström, H, Dorling, P R
• Format: Journal Article
• Language: English
• Published: England 15.11.1983
• Subjects: Acetylglucosaminidase - pharmacology; Alkaloids - pharmacology; Aminopeptidases - biosynthesis; Aminopeptidases - metabolism; Animals; Biological Transport - drug effects; CD13 Antigens; Electrophoresis, Polyacrylamide Gel; Hydrolysis; Intestinal Mucosa - drug effects; Intestinal Mucosa - enzymology; Lectins; Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; Microvilli - drug effects; Microvilli - enzymology; Organ Culture Techniques; Plant Lectins; Protein Processing, Post-Translational - drug effects; Swainsonine; Swine
• ISSN: 0264-6021; 0306-3283; 1470-8728
• DOI: 10.1042/bj2160325

The post-translational processing of pig small-intestinal aminopeptidase N (EC 3.4.11.2) was studied in organ-cultured mucosal explants. Exposure of the explants to swainsonine, an inhibitor of Golgi mannosidase II, resulted in the formation of a Mr-160000 polypeptide, still sensitive to endo-beta-N-acetylglucosaminidase H. Swainsonine caused only a moderate inhibition of transport of the enzyme through the Golgi complex and the subsequent expression in the microvillar membrane. This may imply that the trimming of the high-mannose core and complex glycosylation of N-linked oligosaccharides is not essential for the transport of aminopeptidase N to its final destination. A different type of processing was observed to take place in the presence of swainsonine, resulting in a considerable increase in apparent Mr (from 140000 to 160000). This processing could not be ascribed to N-linked glycosylation, since treatment of the Mr-160000 polypeptide with endo-beta-N-acetylglucosaminidase H only decreased its apparent Mr by 15000. The susceptibility of the mature Mr-166000 polypeptide, but not the Mr-140000 polypeptide, to mild alkaline hydrolysis suggests that aminopeptidase N becomes glycosylated with O-linked oligosaccharides during its passage through the Golgi complex. Aminopeptidase N was not labelled by [3H]palmitic acid, indicating that the processing of the enzyme does not include acylation.