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. 1995 Nov 21;92(24):11044–11048. doi: 10.1073/pnas.92.24.11044

Peptide conjugation to an in vitro-selected DNA ligand improves enzyme inhibition.

Y Lin 1, A Padmapriya 1, K M Morden 1, S D Jayasena 1
PMCID: PMC40567  PMID: 7479933

Abstract

An in vitro selection technique was used to identify a specific high-affinity DNA ligand targeted to human neutrophil elastase (HNE). 1H NMR data and a comparative analysis of the selected sequences suggest that the DNA folds into a G-quartet structure with duplexed ends. The high-affinity binding DNA alone did not inhibit the enzymatic activity of HNE. The DNA was covalently attached to a tetrapeptide, N-methoxysuccinyl-Ala-Ala-Pro-Val, that is a weak competitive inhibitor of HNE. HNE was inhibited by this DNA-peptide conjugate nearly five orders of magnitude more effectively than by the peptide alone. These results demonstrate that in vitro-selected nucleic acids can be used as a vehicle for molecular delivery.

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Selected References

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

  1. Bartel D. P., Szostak J. W. Isolation of new ribozymes from a large pool of random sequences [see comment]. Science. 1993 Sep 10;261(5127):1411–1418. doi: 10.1126/science.7690155. [DOI] [PubMed] [Google Scholar]
  2. Bartel D. P., Zapp M. L., Green M. R., Szostak J. W. HIV-1 Rev regulation involves recognition of non-Watson-Crick base pairs in viral RNA. Cell. 1991 Nov 1;67(3):529–536. doi: 10.1016/0092-8674(91)90527-6. [DOI] [PubMed] [Google Scholar]
  3. Bock L. C., Griffin L. C., Latham J. A., Vermaas E. H., Toole J. J. Selection of single-stranded DNA molecules that bind and inhibit human thrombin. Nature. 1992 Feb 6;355(6360):564–566. doi: 10.1038/355564a0. [DOI] [PubMed] [Google Scholar]
  4. Connell G. J., Illangesekare M., Yarus M. Three small ribooligonucleotides with specific arginine sites. Biochemistry. 1993 Jun 1;32(21):5497–5502. doi: 10.1021/bi00072a002. [DOI] [PubMed] [Google Scholar]
  5. Ellington A. D., Szostak J. W. In vitro selection of RNA molecules that bind specific ligands. Nature. 1990 Aug 30;346(6287):818–822. doi: 10.1038/346818a0. [DOI] [PubMed] [Google Scholar]
  6. Hardin C. C., Henderson E., Watson T., Prosser J. K. Monovalent cation induced structural transitions in telomeric DNAs: G-DNA folding intermediates. Biochemistry. 1991 May 7;30(18):4460–4472. doi: 10.1021/bi00232a013. [DOI] [PubMed] [Google Scholar]
  7. Janda K. D. Tagged versus untagged libraries: methods for the generation and screening of combinatorial chemical libraries. Proc Natl Acad Sci U S A. 1994 Nov 8;91(23):10779–10785. doi: 10.1073/pnas.91.23.10779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Janoff A., Feinstein G., Malemud C. J., Elias J. M. Degradation of cartilage proteoglycan by human leukocyte granule neutral proteases--a model of joint injury. I. Penetration of enzyme into rabbit articular cartilage and release of 35SO4-labeled material from the tissue. J Clin Invest. 1976 Mar;57(3):615–624. doi: 10.1172/JCI108317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Jellinek D., Green L. S., Bell C., Janjić N. Inhibition of receptor binding by high-affinity RNA ligands to vascular endothelial growth factor. Biochemistry. 1994 Aug 30;33(34):10450–10456. doi: 10.1021/bi00200a028. [DOI] [PubMed] [Google Scholar]
  10. Jellinek D., Lynott C. K., Rifkin D. B., Janjić N. High-affinity RNA ligands to basic fibroblast growth factor inhibit receptor binding. Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):11227–11231. doi: 10.1073/pnas.90.23.11227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Jenison R. D., Gill S. C., Pardi A., Polisky B. High-resolution molecular discrimination by RNA. Science. 1994 Mar 11;263(5152):1425–1429. doi: 10.1126/science.7510417. [DOI] [PubMed] [Google Scholar]
  12. Kaplan P. D., Kuhn C., Pierce J. A. The induction of emphysema with elastase. I. The evolution of the lesion and the influence of serum. J Lab Clin Med. 1973 Sep;82(3):349–356. [PubMed] [Google Scholar]
  13. Kubik M. F., Stephens A. W., Schneider D., Marlar R. A., Tasset D. High-affinity RNA ligands to human alpha-thrombin. Nucleic Acids Res. 1994 Jul 11;22(13):2619–2626. doi: 10.1093/nar/22.13.2619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lehman N., Joyce G. F. Evolution in vitro of an RNA enzyme with altered metal dependence. Nature. 1993 Jan 14;361(6408):182–185. doi: 10.1038/361182a0. [DOI] [PubMed] [Google Scholar]
  15. Lin Y., Qiu Q., Gill S. C., Jayasena S. D. Modified RNA sequence pools for in vitro selection. Nucleic Acids Res. 1994 Dec 11;22(24):5229–5234. doi: 10.1093/nar/22.24.5229. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lorsch J. R., Szostak J. W. In vitro evolution of new ribozymes with polynucleotide kinase activity. Nature. 1994 Sep 1;371(6492):31–36. doi: 10.1038/371031a0. [DOI] [PubMed] [Google Scholar]
  17. Lorsch J. R., Szostak J. W. In vitro selection of RNA aptamers specific for cyanocobalamin. Biochemistry. 1994 Feb 1;33(4):973–982. doi: 10.1021/bi00170a016. [DOI] [PubMed] [Google Scholar]
  18. Macaya R. F., Schultze P., Smith F. W., Roe J. A., Feigon J. Thrombin-binding DNA aptamer forms a unimolecular quadruplex structure in solution. Proc Natl Acad Sci U S A. 1993 Apr 15;90(8):3745–3749. doi: 10.1073/pnas.90.8.3745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Nakajima K., Powers J. C., Ashe B. M., Zimmerman M. Mapping the extended substrate binding site of cathepsin G and human leukocyte elastase. Studies with peptide substrates related to the alpha 1-protease inhibitor reactive site. J Biol Chem. 1979 May 25;254(10):4027–4032. [PubMed] [Google Scholar]
  20. Obrist R., Sandberg A. L. In vitro effects of antitumor antibody--chemotactic factor complexes. Clin Immunol Immunopathol. 1982 Oct;25(1):91–102. doi: 10.1016/0090-1229(82)90168-4. [DOI] [PubMed] [Google Scholar]
  21. Pietersz G. A., Kanellos J., Smyth M. J., Zalcberg J., McKenzie I. F. The use of monoclonal antibody conjugates for the diagnosis and treatment of cancer. Immunol Cell Biol. 1987 Apr;65(Pt 2):111–125. doi: 10.1038/icb.1987.13. [DOI] [PubMed] [Google Scholar]
  22. Prudent J. R., Uno T., Schultz P. G. Expanding the scope of RNA catalysis. Science. 1994 Jun 24;264(5167):1924–1927. doi: 10.1126/science.8009223. [DOI] [PubMed] [Google Scholar]
  23. Roberts B. L., Markland W., Ley A. C., Kent R. B., White D. W., Guterman S. K., Ladner R. C. Directed evolution of a protein: selection of potent neutrophil elastase inhibitors displayed on M13 fusion phage. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2429–2433. doi: 10.1073/pnas.89.6.2429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Scaria P. V., Shire S. J., Shafer R. H. Quadruplex structure of d(G3T4G3) stabilized by K+ or Na+ is an asymmetric hairpin dimer. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10336–10340. doi: 10.1073/pnas.89.21.10336. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Schneider D., Tuerk C., Gold L. Selection of high affinity RNA ligands to the bacteriophage R17 coat protein. J Mol Biol. 1992 Dec 5;228(3):862–869. doi: 10.1016/0022-2836(92)90870-p. [DOI] [PubMed] [Google Scholar]
  26. Smith F. W., Feigon J. Quadruplex structure of Oxytricha telomeric DNA oligonucleotides. Nature. 1992 Mar 12;356(6365):164–168. doi: 10.1038/356164a0. [DOI] [PubMed] [Google Scholar]
  27. Trail P. A., Willner D., Lasch S. J., Henderson A. J., Hofstead S., Casazza A. M., Firestone R. A., Hellström I., Hellström K. E. Cure of xenografted human carcinomas by BR96-doxorubicin immunoconjugates. Science. 1993 Jul 9;261(5118):212–215. doi: 10.1126/science.8327892. [DOI] [PubMed] [Google Scholar]
  28. Tuerk C., Gold L. Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science. 1990 Aug 3;249(4968):505–510. doi: 10.1126/science.2200121. [DOI] [PubMed] [Google Scholar]
  29. Tuerk C., MacDougal S., Gold L. RNA pseudoknots that inhibit human immunodeficiency virus type 1 reverse transcriptase. Proc Natl Acad Sci U S A. 1992 Aug 1;89(15):6988–6992. doi: 10.1073/pnas.89.15.6988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Weiland J. E., Davis W. B., Holter J. F., Mohammed J. R., Dorinsky P. M., Gadek J. E. Lung neutrophils in the adult respiratory distress syndrome. Clinical and pathophysiologic significance. Am Rev Respir Dis. 1986 Feb;133(2):218–225. doi: 10.1164/arrd.1986.133.2.218. [DOI] [PubMed] [Google Scholar]
  31. Williamson J. R., Raghuraman M. K., Cech T. R. Monovalent cation-induced structure of telomeric DNA: the G-quartet model. Cell. 1989 Dec 1;59(5):871–880. doi: 10.1016/0092-8674(89)90610-7. [DOI] [PubMed] [Google Scholar]

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