2-Benzamidoethyl Group − A Novel Type of Phosphate Protecting Group for Oligonucleotide Synthesis

A number of 5‘-O-(4,4‘-dimethoxytrityl)thymidine N,N-diisopropylamino phosphoramidites protected at P(III) with derivatives of 2-benzamidoethanol were synthesized and incorporated into synthetic oligonucleotides. Depending on substitution patterns at the alkyl chain, amido group, and phenyl ring, th...

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Bibliographic Details
Published in:Journal of the American Chemical Society 2001-02, Vol.123 (5), p.783-793
Main Authors: Guzaev, Andrei P, Manoharan, Muthiah
Format: Article
Language:English
Subjects:
Base Sequence
Benzamides
Magnetic Resonance Spectroscopy
Oligonucleotides - chemical synthesis
Oligonucleotides - chemistry
Phosphates - chemistry
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Summary:A number of 5‘-O-(4,4‘-dimethoxytrityl)thymidine N,N-diisopropylamino phosphoramidites protected at P(III) with derivatives of 2-benzamidoethanol were synthesized and incorporated into synthetic oligonucleotides. Depending on substitution patterns at the alkyl chain, amido group, and phenyl ring, the time required for removal of these protecting groups using concentrated ammonium hydroxide varied from 48 h at 55 °C to 25 min at 25 °C. Of the 11 groups studied, 2-[N-isopropyl-N- (4-methoxybenzoyl)amino]ethyl- (H) and ω-(thionobenzoylamino)alkyl protections (I and K) were most easily removed. Derivatives of the 2-[N-methyl-N-benzoylamino]ethyl group (E−G) demonstrated moderate stability, but those of the 2-(N-benzoylamino)ethyl group (A−C) were the most stable. For the most reactive group, H, a phosphitylating reagent, bisamidite 60, was synthesized and used in the preparation of four deoxynucleoside phosphoramidites 28 and 65−67, plus the 2‘-O-(2-methoxyethyl)-5-methyluridine phosphoramidite 68. All of these novel building blocks were successfully tested in the preparation of natural, 20-mer oligonucleotides and their phosphorothioate analogues. With the model phosphotriester 37, the mechanism of deprotection was studied and revealed, in the case of group H, a pH-independent formation of the 2-oxazolinium cation 47. Under aqueous conditions, 47 gave 54, which in turn was converted in the presence of ammonia to a number of identified products. It is important to note that none of the products formed was reactive toward the oligonucleotide backbone or nucleic bases. Thus, a general strategy for protection of internucleosidic phosphodiester groups is described, which may also find application in synthetic organic chemistry of phosphorus(III) and (V).
ISSN:0002-7863
1520-5126
DOI:10.1021/ja0016396