Preparation of beta-cells from fetal bovine pancreas: characterization of insulin biosynthetic activity.
Dispersed cell preparations enriched in beta-cells were obtained by collagenase digestion of fetal bovine pancreas and separation by Ficoll gradient centrifugation. These cells actively incorporated [3H]leucine into proinsulin and insulin. Incubation of these cells in the presence of the arginine analogue, L-canavanine, resulted in the inhibition of conversion of newly formed proinsulin to insulin and the appearance of a radioactive component of molecular weight 11,000-12,000. Incorporation of [35S]methionine into this component was detected in the presence of canavanine, an event not observed in control incubation. Canavanine thus induced the formation of a component possessing molecular weight and compositional properties expected for preproinsulin. Further characterization of cellular products by polyacrylamide slab gel electrophoresis in sodium dodecyl sulfate showed a highly labeled band corresponding to molecular weight 18,000-20,000 which might be involved in insulin biosynthesis.
D000818:Animals; D002172:Canavanine / Q000494:pharmacology; D002417:Cattle; D066298:In Vitro Techniques; D007328:Insulin / Q000096:biosynthesis*; D007515:Islets of Langerhans / Q000166:cytology* / Q000378:metabolism; D011384:Proinsulin / Q000096:biosynthesis; D014176:Protein Biosynthesis
D007328:Insulin; D002172:Canavanine; D011384:Proinsulin