Biosynthesis of preprorenin. Studies using whole tissue, a cell-free system, and E. coli containing cDNA inserted at the PstI site of plasmid pBR322.
The biosynthesis of renin as a higher molecular weight 'prorenin' was demonstrated by in vitro incorporation of [35S] methionine into nascent polypeptides of submandibular gland tissue from adult male mice. Immunoprecipitation with anti-renin and electrophoresis identified a Mr 44,500, pI 6.4 prorenin which normally represented 5% of renin-immunoreactive protein in tissue extracts and which was rapidly converted during in vitro labeling into a Mr 40,000, pI 6.2 species. The latter was subsequently processed slowly to forms of Mr 35,500, pI 5.6 and Mr 34,000, pI 5.4. The influence of processing enzymes was then eliminated by synthesizing renin in a cell-free translation system containing rabbit reticulocyte lysate and mRNA isolated from submandibular glands. This yielded an even larger species of Mr 46,000 likely to be 'preprorenin'. A clone bank of mouse submandibular gland cDNA was prepared. This consisted of E. coli RRI transformed with plasmid pBR322 in which cDNA had been inserted at the PstI site so that the bacteria could express the encoded polypeptide. Renin-immunoreactive colonies were identified suggesting the expression of renin-like proteins by a prokaryote. The cDNA which was 700-1000 base pairs big was excised for sequencing.
D000818:Animals; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism*; D004792:Enzyme Precursors / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D008297:Male; D051379:Mice; D008817:Mice, Mutant Strains; D010957:Plasmids; D011061:Poly A / Q000235:genetics; D014176:Protein Biosynthesis; D012313:RNA / Q000235:genetics; D012333:RNA, Messenger; D011817:Rabbits; D012083:Renin / Q000235:genetics*; D012156:Reticulocytes / Q000378:metabolism; D013363:Submandibular Gland / Q000201:enzymology*
D004792:Enzyme Precursors; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D004247:DNA; D012083:Renin