Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1997 Jul 15;25(14):2784–2791. doi: 10.1093/nar/25.14.2784

Site-specific introduction of functional groups into phosphodiester oligodeoxynucleotides and their thermal stability and nuclease-resistance properties.

Y Nomura 1, Y Ueno 1, A Matsuda 1
PMCID: PMC146824  PMID: 9207025

Abstract

We report here the site-specific introduction of functional groups into phosphodiester oligodeoxynucleotides (ODNs). ODNs containing both 5-( N-aminohexyl)-carbamoyl-2'-deoxyuridine (H), which serves as a tether for the further conjugation of functional groups, and 5-(N,N-dimethylaminohexyl)carbamoyl-2'-deoxyuridine (D), which contributes to the thermal stability of the duplex and to the resistance to nucleolytic hydrolysis by nucleases, were synthesized. Functional groups such as folic acid and palmitic acid were site-specifically introduced into the terminus of the aminohexyl-linker of H. The thermal stability and resistance toward nuclease digestion of the modified ODNs were studied. We found that ODNs containing D and H formed stable duplexes with both the complementary DNA and RNA strands even when a bulky functional group such as folic acid, palmitic acid or cholesterol was attached to the terminus of the amino-linker. We also found that ODN analogues which contained D were more resistant to nucleolytic degradation by exo- and endonuclease than the unmodified ODN. Furthermore, duplexes formed by ODNs containing D and the complementary RNA could elicit RNase H activity.

Full Text

The Full Text of this article is available as a PDF (137.6 KB).

Selected References

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

  1. Cohen S. S., McCormick F. P. Polyamines and virus multiplication. Adv Virus Res. 1979;24:331–387. doi: 10.1016/s0065-3527(08)60397-8. [DOI] [PubMed] [Google Scholar]
  2. Cullen B. R. The role of Nef in the replication cycle of the human and simian immunodeficiency viruses. Virology. 1994 Nov 15;205(1):1–6. doi: 10.1006/viro.1994.1613. [DOI] [PubMed] [Google Scholar]
  3. Godard G., Boutorine A. S., Saison-Behmoaras E., Hélène C. Antisense effects of cholesterol-oligodeoxynucleotide conjugates associated with poly(alkylcyanoacrylate) nanoparticles. Eur J Biochem. 1995 Sep 1;232(2):404–410. [PubMed] [Google Scholar]
  4. Hayase Y., Inoue H., Ohtsuka E. Secondary structure in formylmethionine tRNA influences the site-directed cleavage of ribonuclease H using chimeric 2'-O-methyl oligodeoxyribonucleotides. Biochemistry. 1990 Sep 18;29(37):8793–8797. doi: 10.1021/bi00489a041. [DOI] [PubMed] [Google Scholar]
  5. Krieg A. M., Tonkinson J., Matson S., Zhao Q., Saxon M., Zhang L. M., Bhanja U., Yakubov L., Stein C. A. Modification of antisense phosphodiester oligodeoxynucleotides by a 5' cholesteryl moiety increases cellular association and improves efficacy. Proc Natl Acad Sci U S A. 1993 Feb 1;90(3):1048–1052. doi: 10.1073/pnas.90.3.1048. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Kropinski A. M., Bose R. J., Warren R. A. 5-(4-Aminobutylaminomethyl)uracil, an unusual pyrimidine from the deoxyribonucleic acid of bacteriophage phiW-14. Biochemistry. 1973 Jan 2;12(1):151–157. doi: 10.1021/bi00725a025. [DOI] [PubMed] [Google Scholar]
  7. Leamon C. P., Low P. S. Delivery of macromolecules into living cells: a method that exploits folate receptor endocytosis. Proc Natl Acad Sci U S A. 1991 Jul 1;88(13):5572–5576. doi: 10.1073/pnas.88.13.5572. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Letsinger R. L., Zhang G. R., Sun D. K., Ikeuchi T., Sarin P. S. Cholesteryl-conjugated oligonucleotides: synthesis, properties, and activity as inhibitors of replication of human immunodeficiency virus in cell culture. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6553–6556. doi: 10.1073/pnas.86.17.6553. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. MacKellar C., Graham D., Will D. W., Burgess S., Brown T. Synthesis and physical properties of anti-HIV antisense oligonucleotides bearing terminal lipophilic groups. Nucleic Acids Res. 1992 Jul 11;20(13):3411–3417. doi: 10.1093/nar/20.13.3411. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Milligan J. F., Matteucci M. D., Martin J. C. Current concepts in antisense drug design. J Med Chem. 1993 Jul 9;36(14):1923–1937. doi: 10.1021/jm00066a001. [DOI] [PubMed] [Google Scholar]
  11. Mishra R. K., Moreau C., Ramazeilles C., Moreau S., Bonnet J., Toulmé J. J. Improved leishmanicidal effect of phosphorotioate antisense oligonucleotides by LDL-mediated delivery. Biochim Biophys Acta. 1995 Nov 7;1264(2):229–237. doi: 10.1016/0167-4781(95)00145-7. [DOI] [PubMed] [Google Scholar]
  12. Neuhard J., Maltman K. L., Warren R. A. Bacteriophage phi W-14-infected Pseudomonas acidovorans synthesizes hydroxymethyldeoxyuridine triphosphate. J Virol. 1980 May;34(2):347–353. doi: 10.1128/jvi.34.2.347-353.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Polushin N. N., Cohen J. S. Antisense pro-drugs: 5'-ester oligodeoxynucleotides. Nucleic Acids Res. 1994 Dec 11;22(24):5492–5496. doi: 10.1093/nar/22.24.5492. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Rothberg K. G., Ying Y. S., Kolhouse J. F., Kamen B. A., Anderson R. G. The glycophospholipid-linked folate receptor internalizes folate without entering the clathrin-coated pit endocytic pathway. J Cell Biol. 1990 Mar;110(3):637–649. doi: 10.1083/jcb.110.3.637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Shea R. G., Marsters J. C., Bischofberger N. Synthesis, hybridization properties and antiviral activity of lipid-oligodeoxynucleotide conjugates. Nucleic Acids Res. 1990 Jul 11;18(13):3777–3783. doi: 10.1093/nar/18.13.3777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Stein C. A., Pal R., DeVico A. L., Hoke G., Mumbauer S., Kinstler O., Sarngadharan M. G., Letsinger R. L. Mode of action of 5'-linked cholesteryl phosphorothioate oligodeoxynucleotides in inhibiting syncytia formation and infection by HIV-1 and HIV-2 in vitro. Biochemistry. 1991 Mar 5;30(9):2439–2444. doi: 10.1021/bi00223a020. [DOI] [PubMed] [Google Scholar]
  17. Takeda T., Ikeda K., Mizuno Y., Ueda T. Synthesis and properties of deoxyoligonucleotides containing putrescinylthymine (nucleosides and nucleotides. LXXVI). Chem Pharm Bull (Tokyo) 1987 Sep;35(9):3558–3567. doi: 10.1248/cpb.35.3558. [DOI] [PubMed] [Google Scholar]
  18. Uchiyama Y., Miura Y., Inoue H., Ohtsuka E., Ueno Y., Ikehara M., Iwai S. Studies of the interactions between Escherichia coli ribonuclease HI and its substrate. J Mol Biol. 1994 Nov 4;243(4):782–791. doi: 10.1016/0022-2836(94)90047-7. [DOI] [PubMed] [Google Scholar]
  19. Vinogradov S. V., Suzdaltseva Y. G., Kabanov A. V. Block polycationic oligonucleotide derivative: synthesis and inhibition of herpes virus reproduction. Bioconjug Chem. 1996 Jan-Feb;7(1):3–6. doi: 10.1021/bc9500913. [DOI] [PubMed] [Google Scholar]
  20. Wang S., Lee R. J., Cauchon G., Gorenstein D. G., Low P. S. Delivery of antisense oligodeoxyribonucleotides against the human epidermal growth factor receptor into cultured KB cells with liposomes conjugated to folate via polyethylene glycol. Proc Natl Acad Sci U S A. 1995 Apr 11;92(8):3318–3322. doi: 10.1073/pnas.92.8.3318. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Warren R. A. Modified bases in bacteriophage DNAs. Annu Rev Microbiol. 1980;34:137–158. doi: 10.1146/annurev.mi.34.100180.001033. [DOI] [PubMed] [Google Scholar]
  22. Wiberg J. S. Amber mutants of bacteriophage T4 defective in deoxycytidine diphosphatase and deoxycytidine triphosphatase. On the role of 5-hydroxymethylcytosine in bacteriophage deoxyribonucleic acid. J Biol Chem. 1967 Dec 25;242(24):5824–5829. [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES