Posttranslational cleavage of rat intestinal lactase occurs at the luminal side of the brush border membrane.
The intestinal sucrase-isomaltase precursor is cleaved at the brush border membrane by luminal proteases. Whether the lactase precursor also is cleaved by luminal proteases is uncertain. Lactase synthesis and processing was studied in 0- and 15-day-old rats after IP administration of [35S]methionine, and changes in precociously cortisone-induced sucrase-isomaltase were used as an internal control. Mucosal lactase and sucrase-isomaltase were separately immunoprecipitated and analyzed by autoradiography after electrophoresis. In both 0- and 15-day-old rats, mucosal lactase appeared as a 200K lactase precursor band at 30 minutes and as 200K and 225K lactase precursor bands at 60 minutes and was cleaved to form a 130K lactase band 120-240 minutes after labeling; sucrase-isomaltase similarly appeared as 210K and 220K bands at 30-60 minutes and was cleaved to form 140K I and 120K S subunits by 240 minutes in day 15 rats. To determine the role of luminal proteases, intestinal segments were isolated in situ and the luminal contents were flushed 30 minutes after labeling. Unflushed segments were used as controls. Only lactase precursor and sucrase-isomaltase precursor were present 240 minutes after labeling in flushed intestinal segments, but lactase precursor and sucrase-isomaltase precursor were cleaved in unflushed segments. Addition of trypsin or elastase into the lumen of flushed segments resulted in partial cleavage of lactase precursor but not of sucrase-isomaltase precursor. Luminal contents collected from the small intestine of day 15 rats 120 and 240 minutes after labeling showed 35S-labeled 130K and 80K polypeptides in lactase immunoprecipitates. It is concluded that intestinal lactase is synthesized as lactase precursor and transported to brush border membrane and cleaved by luminal proteases, and the amino end polypeptide cleaved from lactase precursor is released into the lumen.
Yeh
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KY
Department of Medicine, St. Luke's-Roosevelt Hospital Center, New York, New York.
Yeh
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Pan
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Holt
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PR
D000818:Animals; D004792:Enzyme Precursors / Q000378:metabolism*; D007421:Intestine, Small / Q000201:enzymology* / Q000648:ultrastructure; D043322:Lactase; D008871:Microvilli / Q000201:enzymology; D010196:Pancreatic Elastase / Q000502:physiology; D011499:Protein Processing, Post-Translational; D051381:Rats; D011919:Rats, Inbred Strains; D013394:Sucrase-Isomaltase Complex / Q000378:metabolism*; D014357:Trypsin / Q000502:physiology; D001616:beta-Galactosidase / Q000378:metabolism*
D004792:Enzyme Precursors; D013394:Sucrase-Isomaltase Complex; D043322:Lactase; D001616:beta-Galactosidase; D010196:Pancreatic Elastase; D014357:Trypsin