| Abstract: |
The derivatization of nucleic acids with selenium is a new and highly promising approach to facilitate their three-dimensional structure determination by X-ray crystallography. Here, we report a comprehensive study on the chemical and enzymatic syntheses of RNAs containing 2′-methylseleno (2′-Se-methyl) nucleoside labels. Our approach includes the first synthesis of an appropriate purine nucleoside phosphoramidite building block. Most importantly, a substantially changed RNA solid-phase synthesis cycle, comprising treatment with threo-1,4-dimercapto-2,3-butanediol (DTT) after the oxidation step, is required for a reliable strand elongation. This novel operation allows for the chemical syntheses of multiple Se-labeled RNAs in sizes that can typically be achieved only for nonmodified RNAs. In combination with enzymatic ligation, biologically important RNA targets become accessible for crystallography. Exemplarily, this has been demonstrated for the Diels-Alder ribozyme and the add adenine riboswitch sequences. We point out that the approach documented here has been the chemical basis for the very recent structure determination of the Diels-Alder ribozyme which represents the first novel RNA fold that has been solved via its Se-derivatives. © 2005 American Chemical Society. |
| Keywords: |
unclassified drug; rna; molecular sequence data; rna synthesis; three dimensional imaging; nucleic acids; base sequence; crystallography, x-ray; purines; selenium; nucleic acid conformation; structure analysis; x ray crystallography; oxidation; diels alder reaction; enzymes; oxidation-reduction; synthesis (chemical); derivatization; nucleotide; chemical modification; ligases; nucleic acid; organophosphorus compounds; adenine; deprotection reaction; methyl group; ribozyme; ribonucleosides; rna, catalytic; oligoribonucleotides; selenium derivative; organoselenium compounds; enzymatic synthases; 1,4 dimercapto 2,3 butanediol; 2,3 butanediol; phosphoramidous acid derivative; purine nucleoside; butylene glycols; dimercaprol
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