Biological activity of rat proinsulin synthesized by Escherichia coli.
Fused proteins possessing insulin antigenic determinants are synthesized and secreted by several strains of Escherichia coli K-12 bearing plasmids containing cDNA copies of rat proinsulin mRNA. Small amounts of the secreted, fused protein from E. coli strains chi 1776 and HB101 bearing plasmid p147 were partially purified and assayed for biological activity by determining stimulation of [1-14C]glucose oxidation to 14CO2 in the rat epididymal fat pad assay. Guinea pig anti-insulin serum (GPAIS) was used to suppress glucose oxidation stimulated by insulin. After mild treatment with trypsin, fused protein stimulated 14CO2 production, and over 90% of this activity was suppressible by GPAIS. Untrypsinized fused protein demonstrated no GPAIS-suppressible 14CO2 production. Larger amounts of fused protein were obtained by transformation of strain PR13 of E. coli in which the yield of eukaryotic proteins is increased approx. 50-fold. A single intravenous injection of trypsinized fused protein from E. coli PR13 containing plasmid p287.47 resulted in a rapid decline in the plasma glucose levels of fasted mice, and the hypoglycemic effect persisted for at least 120 min.
D000273:Adipose Tissue / Q000187:drug effects; D000818:Animals; D001681:Biological Assay; D001786:Blood Glucose / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D008297:Male; D051379:Mice; D010957:Plasmids; D011384:Proinsulin / Q000235:genetics* / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D011917:Rats, Inbred Lew; D014357:Trypsin
D001786:Blood Glucose; D012333:RNA, Messenger; D004247:DNA; D011384:Proinsulin; D014357:Trypsin