Solid support synthesis of all-Rp-oligo(ribonucleoside phosphorothioate)s

被引:28
作者
Almer, H [1 ]
Stawinski, J [1 ]
Stromberg, R [1 ]
机构
[1] UNIV STOCKHOLM, ARRHENIUS LAB, DEPT ORGAN CHEM, S-10691 STOCKHOLM, SWEDEN
关键词
D O I
10.1093/nar/24.19.3811
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The first method for solid support synthesis of all-R(P)-oligo(ribonucleoside phosphorothioate)s is presented as well as attempts to increase the stereoselectivity of the key step in this approach, The synthetic strategy consists of (i) a solid support synthesis procedure, using 5'-O-(4-methoxytriphenylmethyl)-2'-O-tert-butyldimethylsilyl-ribonucleoside 3'-H-phosphonates, that due to stereoselectivity in the condensation step, gives oligomers with mostly S-P-H-phosphonate diesters (72-89% under standard conditions), (ii) stereospecific sulfurization with S-8 in pyridine to produce oligo(ribonucleoside phophorothioate)s enriched with internucleosidic linkages of R(P) configuration, (iii) treatment of the deprotected oligonucleotides with the enzyme Nuclease P1 from Penicillium citrinum, that specifically catalyses cleavage of S-P-phosphorothioate diester linkages, which leaves a mixture of oligomers having all internucleosidic linkages as R(P)-phosphorothioates, and finally (iv) isolation and HPLC purification of the full length all-R(P) oligomer, Mixed sequences containing the four common nucleosidic residues up to the chain length of a heptamer were synthesized, Change of N-4-protection on the cytidine building block from propionyl to N-methylpyrrolidin-2-ylidene gave a slightly improved diastereoselectivity in H-phosphonate diester formation, Increased selectivity up to 99+% was obtained with the guanosine building block when the amount of pyridine in the coupling step was reduced.
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收藏
页码:3811 / 3820
页数:10
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