Preliminary studies were conducted to develop a cell-free system for insulin biosynthesis using the streptozotocin-nicotinamide-induced rat islet adenoma. Radiolabeled proteins, migrating on steric exclusion chromatography and SDS-gel electrophoresis in the region of insulin and proinsulin, were synthesized in a system prepared from the tumor 800 X g supernatant fraction, rat liver cytosol, and appropriate energy substrates. The proteins were not synthesized by a rat liver cell-free system, and synthesis could be substantially inhibited by the addition of cycloheximide. In addition, it could be shown that the islet proteins were not the products of residual intact cells within the system, nor were they an artifact due to nonspecific binding of [3H]-L-leucine to pre-existing insulin and proinsulin. The radiolabeled material eluting with insulin on steric exclusion chromatography was identified as [3H]-insulin by immunoaffinity column chromatography.
Bibliographical noteFunding Information:
From the Departments of Pediatrics and Anatomy, University of Minnesota Medical School, Minneapolis, Minn. Supported in part by grants from the USPHS (AM 24090) and the Diabetes Research Fund. Inc. Address reprint requests to Dana E. Johnson, MD. PhD, University of Minnesota Hospitals, Box 211 Mayo Memorial Hospital, 420 Delaware St SE, Minneapolis, MN 55455. 0 I984 by Grune & Stratton, Inc. 0026-0495/84/3301~002%01.00/0