Mapping of T7 RNA polymerase active site with novel reagents – oligonucleotides with reactive dialdehyde groups

AbstractOligonucleotides of a novel type containing 2′-O-β-ribofuranosyl-cytidine were synthesized and further oxidized to yield T7 consensus promoters with dialdehyde groups. Both types of oligonucleotides were tested as templates, inhibitors, and affinity reagents for T7 RNA polymerase and its mutants. All oligonucleotides tested retained high affinity towards the enzyme. Wild-type T7 RNA polymerase and most of the mutants did not react irreversibly with oxidized oligonucleotides. Affinity labeling was observed only with the promoter-containing dialdehyde group in position (+2) of the coding chain and one of the mutants tested, namely Y639K. These results allowed us to propose the close proximity of residue 639 and the initiation region of the promoter within initiation complex. We suggest the oligonucleotides so modified may be of general value for the study of protein-nucleic acid interactions

Disaccharide nucleosides, an important group of natural compounds

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Preparation of pyrimidine 5’-O-beta-D-ribofuranosyl nucleosides. Hydrolytic stability of O-D-ribofuranosyl nucleosides

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Oligodeoxynucleotides Containing N1-Methyl-2′-Deoxyadenosine and N6-Methyl-2′-Deoxyadenosine

This unit describes a simple and efficient synthesis of the phosphoramidite derivative of N(1)-methyl-2'-deoxyadenosine from 2'-deoxyadenosine. The synthesis starts with the monomethoxytritylation of 2'-deoxyadenosine followed by methylation of 5'-O-protected nucleoside at N-1. Subsequent N-chloroacetylation leads to N(6)-chloroacetyl-N(1)-methyl-5'-O-(p-anisyldiphenylmethyl)-2'-deoxyadenosine, which is finally converted to its 3' phosphoramidite derivative. This phosphoramidite is used to incorporate N(1)-methyl-2'-deoxyadenosine into synthetic oligonucleotides. N-Chloroacetyl protection and controlled anhydrous deprotection conditions are used to avoid the Dimroth rearrangement.status: publishe

Protection of 1-methyladenosine and its chemical incorporation into oligonucleotides

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Chemical incorporation of 1-methyladenosine into oligonucleotides

The base moiety of 1-N-methyladenosine can be protected with a chloroacetyl group for incorporation of this modified nucleoside into DNA and RNA. Carefully controlled anhydrous conditions are needed for deprotection of the oligonucleotides

Chemical incorporation of 1-methyladenosine into oligonucleotides

The base moiety of 1-N-methyladenosine can be protected with a chloroacetyl group for incorporation of this modified nucleoside into DNA and RNA. Carefully controlled anhydrous conditions are needed for deprotection of the oligonucleotides.status: publishe

Chemical incorporation of minor tRNA component O-beta-D-ribofuranosyl-(1"-2')-adenosine-5"-phosphate into oligoribonucleotides

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Solution structure of a RNA decamer duplex, containing 9-[2-O-(beta-D-ribofuranosyl)-beta-D-ribofuranosyl]adenine, a special residue in lower Eukaryotic initiator tRNAs

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