20 research outputs found
Synthesis of Phosphorothioate Oligonucleotides with Stereodefined Phosphorothioate Linkages
A method for solidâphase synthesis of stereodefined PSâoligos via an oxathiaphospholane approach using pure Pâdiastereomers of nucleoside oxathiaphospholane monomers is described. The oxathiaphospholane monomers are synthesized by phosphitylation of 5â˛âOâDMTrâNâprotected deoxyribonucleosides with 2âchloroâspiroâ4,4âpentamethyleneâ1,3,2âoxathiaphospholane followed by sulfurization. The procedure is general and may be applied to other analogs, depending on the aldehyde (or mercaptoalcohol) used. Starting from an 18Oâlabeled mercaptoalcohol, the corresponding 18Oâlabeled phosphitylating reagent and nucleoside monomers can be obtained and used for synthesis of labeled stereodefined PSâoligos, which are useful for studying mechanisms of enzymatic reactions. Details are provided for chromatographic separation of the 5â˛âOâDMTrâNâprotectedâdeoxyribonucleosideâ3â˛âOâ(2âthioâspiroâ4,4âpentamethyleneâ1,3,2âoxathiaphospholane)s into their Pâdiastereomers, and for manual solidâphase synthesis of PSâoligos. Oxidation of 5â˛âOâDMTrâNâprotectedâdeoxyribonucleosideâ3â˛âOâ(2âthioâspiroâ4,4âpentamethyleneâ1,3,2âoxathiaphospholane)s with selenium dioxide yields their 2âoxoâanalogs, which are suitable either for elongation of stereodefined PSâoligos with segments consisting of unmodified nucleotide units possessing phosphate internucleotide linkages, or for generating isotopomeric 18Oâlabeled POâoligos of predetermined Pâchirality.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/143684/1/cpnc0417.pd
Structure of a stereoregular phosphorothioate DNA/RNA duplex
In this work, we present the first NMR solution structure of a DNA/RNA hybrid containing stereoregular Rp-phosphorothioate modifications of all DNA backbone linkages. The complex of the enzymatically synthesized phosphorothioate DNA octamer (all-R_p)-d(GCGTCAGG) and its complementary RNA r(CCUGACGC) was found to adopt an overall conformation within the A-form family. Most helical parameters and the sugar puckers of the DNA strand assume values intermediate between A- and B-form. The close structural similarity with the unmodified DNA/RNA hybrid of the same sequence may explain why both the natural and the sulfur-substituted complex can be recognized and digested by ribonuclease H