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. 1979 Jun;76(6):2689–2693. doi: 10.1073/pnas.76.6.2689

De novo biosynthesis of an enzymatically active precursor form of bovine pancreatic RNase.

T H Haugen, E C Heath
PMCID: PMC383673  PMID: 288056

Abstract

The de novo biosynthesis of RNase (ribonucleate 3'-pyrimidino-oligonucleotidohydrolase, EC 3.1.4.22) was studied in a cell-free rabbit reticulocyte translation system using a polyadenylylated fraction of mRNA isolated from bovine pancreas. Analysis of the [35S]methionine-labeled translation products of pancreas mRNA by polyacrylamide gel electrophoresis indicated the presence of several proteins, one of which corresponded to 16,500 daltons, or approximately 2800 daltons greater than native RNase A. This protein was specifically precipitated from the mixture of products by anti-RNase antibodies. Partial sequence determination of the NH2-terminal region of the anti-RNase antibody-precipitable species indicated that it is a precursor form of RNase A with 25 additional amino acids on its NH2 terminus. The precursor nature of the protein was confirmed by demonstration that a mixture of RNase A and a glycosylated form of the enzyme, RNase B, is formed when translation of the mRNA is conducted in the presence of dog pancreas membranes. Assay of the putative precursor form of RNase for catalytic activity with polycytidylic acid as substrate indicated that the protein has a specific enzymatic activity identical to that of native RNase A.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. ANFINSEN C. B., HABER E. Studies on the reduction and re-formation of protein disulfide bonds. J Biol Chem. 1961 May;236:1361–1363. [PubMed] [Google Scholar]
  2. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blackburn P., Wilson G., Moore S. Ribonuclease inhibitor from human placenta. Purification and properties. J Biol Chem. 1977 Aug 25;252(16):5904–5910. [PubMed] [Google Scholar]
  4. Blobel G., Dobberstein B. Transfer of proteins across membranes. II. Reconstitution of functional rough microsomes from heterologous components. J Cell Biol. 1975 Dec;67(3):852–862. doi: 10.1083/jcb.67.3.852. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Katz F. N., Rothman J. E., Lingappa V. R., Blobel G., Lodish H. F. Membrane assembly in vitro: synthesis, glycosylation, and asymmetric insertion of a transmembrane protein. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3278–3282. doi: 10.1073/pnas.74.8.3278. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  7. Lomedico P. T., Chan S. J., Steiner D. F., Saunders G. F. Immunological and chemical characterization of bovine preproinsulin. J Biol Chem. 1977 Nov 25;252(22):7971–7978. [PubMed] [Google Scholar]
  8. MacDonald R. J., Przybyla A. E., Rutter W. J. Isolation and in vitro translation of the messenger RNA coding for pancreatic amylase. J Biol Chem. 1977 Aug 10;252(15):5522–5528. [PubMed] [Google Scholar]
  9. PLUMMER T. H., Jr, HIRS C. H. The isolation of ribounclease B, a glycoprotein, from bovine pancreatic juice. J Biol Chem. 1963 Apr;238:1396–1401. [PubMed] [Google Scholar]
  10. Palacios R., Palmiter R. D., Schimke R. T. Identification and isolation of ovalbumin-synthesizing polysomes. I. Specific binding of 125 I-anti-ovalbumin to polysomes. J Biol Chem. 1972 Apr 25;247(8):2316–2321. [PubMed] [Google Scholar]
  11. Pelham H. R., Jackson R. J. An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976 Aug 1;67(1):247–256. doi: 10.1111/j.1432-1033.1976.tb10656.x. [DOI] [PubMed] [Google Scholar]
  12. Plummer T. H., Jr Glycoproteins of bovine pancreatic juice. Isolation of ribonucleases C and D. J Biol Chem. 1968 Nov 25;243(22):5961–5966. [PubMed] [Google Scholar]
  13. Stark N. J., Heath E. C. Glucose-dependent glycosylation of secretory glycoprotein in mouse myeloma cells. Arch Biochem Biophys. 1979 Feb;192(2):599–609. doi: 10.1016/0003-9861(79)90131-0. [DOI] [PubMed] [Google Scholar]
  14. Ullrich A., Shine J., Chirgwin J., Pictet R., Tischer E., Rutter W. J., Goodman H. M. Rat insulin genes: construction of plasmids containing the coding sequences. Science. 1977 Jun 17;196(4296):1313–1319. doi: 10.1126/science.325648. [DOI] [PubMed] [Google Scholar]

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