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. 1981 Jun;78(6):3313–3317. doi: 10.1073/pnas.78.6.3313

Expression in Escherichia coli of biologically active enzyme by a DNA sequence coding for the human plasminogen activator urokinase.

B Ratzkin, S G Lee, W J Schrenk, R Roychoudhury, M Chen, T A Hamilton, P P Hung
PMCID: PMC319558  PMID: 7022447

Abstract

We have isolated clones of Escherichia coli strain K-12 that contain a hybrid pBR322 plasmid having a 4.2-kilobase insert of a DNA transcript of the mRNA of human plasminogen activator, urokinase. The bacterially produced enzyme has properties similar to those of urokinase from human fetal kidney cells. Both enzymes occur in discrete forms ranging from 32,000 to 150,000 daltons in size. They react with antibody to purified urokinase from human kidney cells, bind to a benzamidine-Sepharose column, and induce plasminogen-dependent lysis of a fibrin clot.

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Selected References

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

  1. Bernik M. B., Kwaan H. C. Origin of fibrinolytic activity in cultures of the human kidney. J Lab Clin Med. 1967 Oct;70(4):650–661. [PubMed] [Google Scholar]
  2. Clarke L., Hitzeman R., Carbon J. Selection of specific clones from colony banks by screening with radioactive antibody. Methods Enzymol. 1979;68:436–442. doi: 10.1016/0076-6879(79)68033-3. [DOI] [PubMed] [Google Scholar]
  3. Deeley R. G., Gordon J. I., Burns A. T., Mullinix K. P., Binastein M., Goldberg R. F. Primary activation of the vitellogenin gene in the rooster. J Biol Chem. 1977 Nov 25;252(22):8310–8319. [PubMed] [Google Scholar]
  4. Friedman E. Y., Rosbash M. The syntheiss of high yields of full-length reverse transcripts of globin mRNA. Nucleic Acids Res. 1977 Oct;4(10):3455–3471. doi: 10.1093/nar/4.10.3455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Goeddel D. V., Heyneker H. L., Hozumi T., Arentzen R., Itakura K., Yansura D. G., Ross M. J., Miozzari G., Crea R., Seeburg P. H. Direct expression in Escherichia coli of a DNA sequence coding for human growth hormone. Nature. 1979 Oct 18;281(5732):544–548. doi: 10.1038/281544a0. [DOI] [PubMed] [Google Scholar]
  6. Holmberg L., Bladh B., Astedt B. Purification of urokinase by affinity chromatography. Biochim Biophys Acta. 1976 Aug 12;445(1):215–222. doi: 10.1016/0005-2744(76)90174-1. [DOI] [PubMed] [Google Scholar]
  7. Jacobsen H., Klenow H., Overgaard-Hansen K. The N-terminal amino-acid sequences of DNA polymerase I from Escherichia coli and of the large and the small fragments obtained by a limited proteolysis. Eur J Biochem. 1974 Jun 15;45(2):623–627. doi: 10.1111/j.1432-1033.1974.tb03588.x. [DOI] [PubMed] [Google Scholar]
  8. Kessler S. W. Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975 Dec;115(6):1617–1624. [PubMed] [Google Scholar]
  9. LESUK A., TERMINIELLO L., TRAVER J. H. CRYSTALLINE HUMAN UROKINASE: SOME PROPERTIES. Science. 1965 Feb 19;147(3660):880–882. doi: 10.1126/science.147.3660.880. [DOI] [PubMed] [Google Scholar]
  10. 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]
  11. Ling C. M., Overby L. R. Prevalence of hepatitis B virus antigen as revealed by direct radioimmune assay with 125 I-antibody. J Immunol. 1972 Oct;109(4):834–841. [PubMed] [Google Scholar]
  12. Livingston D. M. Immunoaffinity chromatography of proteins. Methods Enzymol. 1974;34:723–731. doi: 10.1016/s0076-6879(74)34094-3. [DOI] [PubMed] [Google Scholar]
  13. Meagher R. B., Tait R. C., Betlach M., Boyer H. W. Protein expression in E. coli minicells by recombinant plasmids. Cell. 1977 Mar;10(3):521–536. doi: 10.1016/0092-8674(77)90039-3. [DOI] [PubMed] [Google Scholar]
  14. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  15. 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]
  16. Ratzkin B., Carbon J. Functional expression of cloned yeast DNA in Escherichia coli. Proc Natl Acad Sci U S A. 1977 Feb;74(2):487–491. doi: 10.1073/pnas.74.2.487. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rhoads R. E., McKnight G. S., Schimke R. T. Quantitative measurement of ovalbumin messenger ribonucleic acid activity. Localization in polysomes, induction by estrogen, and effect of actinomycin D. J Biol Chem. 1973 Mar 25;248(6):2031–2039. [PubMed] [Google Scholar]
  18. Roberts B. E., Paterson B. M. Efficient translation of tobacco mosaic virus RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2330–2334. doi: 10.1073/pnas.70.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Roychoudhury R., Jay E., Wu R. Terminal labeling and addition of homopolymer tracts to duplex DNA fragments by terminal deoxynucleotidyl transferase. Nucleic Acids Res. 1976 Apr;3(4):863–877. doi: 10.1093/nar/3.4.863. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Seeburg P. H., Shine J., Martial J. A., Ivarie R. D., Morris J. A., Ullrich A., Baxter J. D., Goodman H. M. Synthesis of growth hormone by bacteria. Nature. 1978 Dec 21;276(5690):795–798. doi: 10.1038/276795a0. [DOI] [PubMed] [Google Scholar]
  21. Unkeless J., Dano K., Kellerman G. M., Reich E. Fibrinolysis associated with oncogenic transformation. Partial purification and characterization of the cell factor, a plasminogen activator. J Biol Chem. 1974 Jul 10;249(13):4295–4305. [PubMed] [Google Scholar]
  22. WILLIAMS J. R. B. The fibrinolytic activity of urine. Br J Exp Pathol. 1951 Dec;32(6):530–537. [PMC free article] [PubMed] [Google Scholar]
  23. White W. F., Barlow G. H., Mozen M. M. The isolation and characterization of plasminogen activators (urokinase) from human urine. Biochemistry. 1966 Jul;5(7):2160–2169. doi: 10.1021/bi00871a003. [DOI] [PubMed] [Google Scholar]

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