Oligodeoxynucleotides containing unnatural L-2′-deoxyribose

Damha, Masad J.; Giannaris, Paul A.; Marfey, Peter S.; Reid, Lorne S.

doi:10.1016/s0040-4039(00)78788-0

Cited by 39 publications

(28 citation statements)

References 5 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We tested the L-and D-aptamers using purified S1 nuclease (a single-stranded endonuclease) and DNase I (a double-stranded endonuclease) and found that the L-aptamer was unaffected after incubation for 10 days with either nuclease under conditions in which the D-aptamer wasd degraded in under 10 s (see Materials and Methods). Others have performed similar tests with S1 nuclease (15,31), P1 nuclease (a single-stranded endonuclease) (15,30), spleen phosphodiesterase (a 5Ј exonuclease) (15,31,32), and venom phosphodiesterase (a 3Ј exonuclease) (15,33). Of these, only venom phosphodiesterase was found to degrade L-DNA; however, hydrolysis was 10,000 times slower than for D-DNA (15,33).…”

Section: Resultsmentioning

confidence: 99%

“…The four L-deoxynucleoside phosphoramidites and L-dA resin were custom-synthesized by Chem-Genes (Waltham, MA). Because it would not otherwise be a substrate for T4 polynucleotide kinase (15), one preparation of L-aptamer contained a 5Ј tail of two D-thymidine residues; this modification had no detectable effect on binding activity. The control L-DNA differed from the L-DNA aptamer in two segments, which had reversed 5Ј to 3Ј polarity [sequence 5Ј-T-(54-26)-(19-25)-(18-2)-A; numbers refer to segments of the aptamer of Fig.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Bioactive and nuclease-resistant l -DNA ligand of vasopressin

Williams

Liu

Schumacher

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

199

128

View full text Add to dashboard Cite

In vitro selection experiments have produced nucleic acid ligands (aptamers) that bind tightly and specifically to a great variety of target biomolecules. The utility of aptamers is often limited by their vulnerability to nucleases present in biological materials. One way to circumvent this problem is to select an aptamer that binds the enantiomer of the target, then synthesize the enantiomer of the aptamer as a nuclease-insensitive ligand of the normal target. We have so identified a mirror-image single-stranded DNA that binds the peptide hormone vasopressin and have demonstrated its stability to nucleases and its bioactivity as a vasopressin antagonist in cell culture.The enantiomer of an autogenously folding macromolecule will assume the mirror-image structure of the parent molecule, as has been documented for proteins and nucleic acids (1, 2). The corollary that the enantiomers of two complex-forming molecules will interact identically also has been demonstrated, for example by inverting all chiral centers in both an enzyme and its substrate (3). The principle has application to a problem in biotechnology, namely, that the peptide or nucleic acid ligands generated by phage display or in vitro selection are susceptible to enzymatic degradation; for example, a DNA oligonucleotide injected i.v. is degraded with a half-life of Ϸ5 min (4-7). The principle of chiral inversion can be coupled to the powerful reiterable selection methods to generate ligands that are insusceptible to degradative enzymes. First, a peptide or nucleic acid ligand is generated that binds the synthetic enantiomer of the target molecule. Then, the enantiomer of the selected ligand is synthesized from D amino acids or L nucleotides. The resulting reflected ligand will then bind and possibly inhibit the target molecule ( Fig. 1). This ''selectionreflection'' strategy has been used recently in the identification of a D peptide ligand of a Src homology 3 domain (8) and L-RNA ligands of arginine and adenosine (9, 10).In our study, the vertebrate hormone arginine vasopressin (VP), a 9-residue cyclic L peptide, was chosen as a target for its ease of bioassay, its ease of synthesis in all-D form, and its cyclic structure that restricts conformational degrees of freedom relative to a linear peptide and may thus allow selection of a tighter binding ligand. A stable ligand of VP could be useful as a diagnostic reagent or as a therapeutic antagonist in diseases associated with excessive levels of VP in blood or cerebrospinal fluid. DNA was chosen as ligand material because much higher pool diversity can be achieved than for peptide selection, and the chemical stability of DNA is superior to that of RNA.We have identified a mirror image DNA that binds VP and have shown that this ligand is bioactive, specifically antagonizing the VP response of cultured kidney cells. Selectionreflection thus joins the approach of using modified RNA polymerase substrates (11-14) as a viable means of coupling the power of in vitro selection to the production of sta...

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Bioactive and nuclease-resistant l -DNA ligand of vasopressin

Williams

Liu

Schumacher

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

199

128

View full text Add to dashboard Cite

show abstract

“…20,23,24 To consider the reason of this discrepancy from the viewpoints of the structural features, molecular modeling was carried out for duplexes [1] and [2]. Energy-minimized structures of the duplexes are shown in Fig.…”

Section: Thermodynamics Of L/d-d Duplexesmentioning

confidence: 99%

“…A homo hexamer of Ldeoxyadenosine, L-(dA)6, bound to poly(U), but not to poly(A), poly(G), poly(C), or poly(dT). 17 Similarly, L-(U)12 and L-(dU) 20 bound to poly(A), but not to poly(dA). 18 In both cases, Loligomers distinguished RNA from DNA.…”

mentioning

confidence: 97%

“…19 Only the chimeric oligomers with both D-and L-2′-deoxyribose sugars (L/D-DNA) achieved both nuclease resistance and duplex formation, though some decrease of duplex stability was found. [20][21][22] NMR studies for duplexes composed of L/D-DNA and complementary natural DNA (L/D-D duplexes) in which single nucleotide was substituted by L-deoxyguanosine (L-dG) or L-thymidine (L-T) revealed that the structures of the duplexes were normal B-form, and the introduced L-nucleotide formed Watson-Crick hydrogen bonds as normal nucleotides. 23,24 However, these findings do not account for the destabilization of duplexes in the other experiments.…”

mentioning

confidence: 99%

See 1 more Smart Citation