ja038481k_si_001.pdf (980.42 kB)
Chemical Synthesis of Selenium-Modified Oligoribonucleotides and Their Enzymatic Ligation Leading to an U6 SnRNA Stem−Loop Segment
journal contribution
posted on 2004-02-04, 00:00 authored by Claudia Höbartner, Ronald MicuraThe derivatization of nucleic acids with selenium is highly promising to facilitate nucleic acids
structure determination by X-ray crystallography using the multiwavelength anomalous dispersion (MAD)
technique. The foundation for such an approach has been laid by Huang, Egli, and co-workers and was
exemplified on small DNA duplexes. Here, we present a comprehensive study on the preparation of RNAs
containing 2‘-Se-methylpyrimidine nucleoside labels. This includes the synthesis of a novel 2‘-Se-methylcytidine phosphoramidite 11 and its incorporation into oligoribonucleotides by solid-phase synthesis.
Deprotection of the oligonucleotides is achieved in the presence of millimolar amounts of threo-1,4-dimercapto-2,3-butandiol (DTT). With this additive, oxidation products and follow-up side-products are
suppressed and acceptable HPLC traces of the crude material are obtained, so far tested for sequences
of up to 22-mers. Moreover, an extensive investigation on the enzymatic ligation of the selenium-containing
oligoribonucleotides demonstrates the high flexibility of the selenium approach. Our target sequence, an
U6 snRNA stem−loop motif comprising all naturally occurring nucleoside modifications beside the Se-label is achieved by ligation using T4 RNA ligase.
History
Usage metrics
Categories
Keywords
selenium approachHPLC tracesoxidation productsMADmillimolar amountsDTTmethylcytidine phosphoramidite 11U 6 snRNAsynthesisligationacids structure determinationoligoribonucleotideSetarget sequencemethylpyrimidine nucleoside labelsEnzymatic Ligationnucleoside modificationsDNA duplexesT 4 RNA ligasechemical Synthesiscrude material
Licence
Exports
RefWorks
BibTeX
Ref. manager
Endnote
DataCite
NLM
DC