Biosynthesis of intestinal microvillar proteins. Processing of N-linked carbohydrate is not required for surface expression.
Castanospermine, an inhibitor of glucosidase I, the initial enzyme in the trimming of N-linked carbohydrate, was used to study the importance of carbohydrate processing in the biosynthesis of microvillar enzymes in organ-cultured pig intestinal explants. For aminopeptidase N (EC 3.4.11.2), aminopeptidase A (EC 3.4.11.7), sucrase-isomaltase (EC 3.2.1.48-10) and maltase-glucoamylase (EC 3.2.1.20), castanospermine caused the formation of novel transient forms of higher Mr than corresponding controls, indicating a blocked removal of glucose residues. For the first three enzymes, the 'mature' (Golgi-processed) forms were similar in size to or slightly smaller than corresponding controls and were, as shown for aminopeptidase N, endoglycosidase-H-sensitive, evidence of a blocked attachment of complex sugars. Maltase-glucoamylase did not undergo conversion into a 'mature' form, suggesting that, unlike other microvillar enzymes, it does not receive post-translational O-linked carbohydrate. Castanospermine suppressed the synthesis of the four enzymes, but did not block their transport to the microvillar membrane, showing that processing of N-linked carbohydrate is not required for microvillar expression. The proteinase inhibitor leupeptin partially restored the suppressed synthesis, indicating that the majority of the wrongly processed enzymes, probably because of conformational instability, become degraded soon after synthesis rather than being transported to the microvillar membrane.
D000470:Alkaloids / Q000494:pharmacology; D000626:Aminopeptidases / Q000378:metabolism; D000818:Animals; D018826:CD13 Antigens; D050260:Carbohydrate Metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D007212:Indolizines; D007413:Intestinal Mucosa / Q000187:drug effects / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology; D008565:Membrane Proteins / Q000096:biosynthesis*; D008871:Microvilli / Q000187:drug effects / Q000378:metabolism; D009924:Organ Culture Techniques; D013552:Swine; D054836:Indolizidines
D000470:Alkaloids; D000626:Aminopeptidases; D018826:CD13 Antigens; D007212:Indolizines; D007976:Leupeptins; D008565:Membrane Proteins; C032854:leupeptin; C037806:castanospermine; D054836:Indolizidines