Journal
CHEMBIOCHEM
Volume 12, Issue 12, Pages 1867-1879Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cbic.201100160
Keywords
DNA; HIV-1 reverse transcriptase; iminodipropionic acid; nucleotides; phosphoramidates
Funding
- K.U. Leuven
- Bijzonder Onderzoeksfonds
- Geconcerteerde Onderzoeksacties
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Previous studies have demonstrated that some selected amino monoacids and amino diacids can function as leaving groups in the polymerase-catalyzed incorporation of deoxynucleotides into DNA. Among these, the iminodiacetic acid phosphoramidate of deoxyadenosine monophosphate (IDA-dAMP) represents an interesting example, as it could overcome some of the problems observed when using l-aspartic acid as the leaving group, that is, poor chain elongation. We have now synthesized and evaluated a series of IDA-dAMP analogues that bear either an extended aliphatic chain in the amino acid function, or a phosphonic acid moiety (substituting for the carboxylic acid function). Among these compounds, the nucleotide with an iminodipropionic acid leaving group (IDP-dAMP) was identified as the best substrate; the excellent single incorporation (91% conversion to a P+1 strand at 50 mu m) was at a substrate concentration ten times lower than that used for IDA-dAMP). This nucleotide also presented improved kinetics and elongation capability compared to IDA-dAMP. The analogues with T, G, and C base moieties were also investigated for their incorporation ability with HIV-1 RT. The incorporation efficiency was found to decrease in the order A > T > G > C. The properties of the iminodipropionic acid as the leaving group surpass those of previously evaluated leaving groups; this acid will be a prime candidate for in vivo testing.
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