Synthesis of phosphorothioate oligonucleotides with stereodefined phosphorothioate linkages.
A method for solid-phase synthesis of stereodefined PS-oligos via an oxathiaphospholane approach using pure P-diastereomers of nucleoside oxathiaphospholane monomers is described. The oxathiaphospholane monomers are synthesized by phosphitylation of 5'-O-DMTr-N-protected deoxyribonucleosides with 2-chloro-spiro-4,4-pentamethylene-1,3,2-oxathiaphospholane followed by sulfurization. The procedure is general and may be applied to other analogs, depending on the aldehyde (or mercaptoalcohol) used. Starting from an 18O-labeled mercaptoalcohol, the corresponding 18O-labeled phosphitylating reagent and nucleoside monomers can be obtained and used for synthesis of labeled stereodefined PS-oligos, which are useful for studying mechanisms of enzymatic reactions. Details are provided for chromatographic separation of the 5'-O-DMTr-N-protected-deoxyribonucleoside-3'-O-(2-thio-spiro-4,4-pentamethylene-1,3,2-oxathiaphospholane)s into their P-diastereomers, and for manual solid-phase synthesis of PS-oligos. Oxidation of 5'-O-DMTr-N-protected-deoxyribonucleoside-3'-O-(2-thio-spiro-4,4-pentamethylene-1,3,2-oxathiaphospholane)s with selenium dioxide yields their 2-oxo-analogs, which are suitable either for elongation of stereodefined PS-oligos with segments consisting of unmodified nucleotide units possessing phosphate internucleotide linkages, or for generating isotopomeric 18O-labeled PO-oligos of predetermined P-chirality.
Guga
Piotr
P
Polish Academy of Sciences, Lodz, Poland.
Stec
Wojciech J
WJ
D002625:Chemistry, Organic / Q000295:instrumentation / Q000379:methods; D003509:Cyclohexanecarboxylic Acids / Q000138:chemical synthesis; D003510:Cyclohexanes / Q000737:chemistry; D004220:Disulfides / Q000138:chemical synthesis; D008954:Models, Biological; D009690:Nucleic Acid Conformation; D009711:Nucleotides / Q000138:chemical synthesis / Q000737:chemistry; D009946:Organothiophosphorus Compounds / Q000138:chemical synthesis; D010710:Phosphates / Q000737:chemistry* / Q000378:metabolism; D054735:Phosphorothioate Oligonucleotides / Q000138:chemical synthesis* / Q000737:chemistry*; D012266:Ribose / Q000031:analogs & derivatives / Q000138:chemical synthesis / Q000737:chemistry; D012521:Sarcosine / Q000737:chemistry; D012997:Solvents / Q000737:chemistry / Q000494:pharmacology; D013237:Stereoisomerism; D019802:Succinic Acid / Q000737:chemistry
D003509:Cyclohexanecarboxylic Acids; D003510:Cyclohexanes; D004220:Disulfides; D009711:Nucleotides; D009946:Organothiophosphorus Compounds; D010710:Phosphates; D054735:Phosphorothioate Oligonucleotides; D012997:Solvents; C434797:oxathiaphospholane; C506365:Cyclohexane; D012266:Ribose; C083120:phosphorodithioic acid; D019802:Succinic Acid; D012521:Sarcosine