期刊
NUCLEIC ACIDS RESEARCH
卷 47, 期 10, 页码 4927-4939出版社
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkz252
关键词
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资金
- Fonds Wetenschappelijk Onderzoek (FWO, Flanders Research Foundation) [G. 078014N, 12Q8619N]
- Research Fund KU Leuven [OT/14/128]
- European Research Council under the European Union's Seventh Framework Program (FP7/2007-2013)/ERC [ERC-2012-ADG 20120216/320683]
- FWO [12Q8619N, G.078014N]
Biomedical applications of nucleic acid aptamers are limited by their rapid degradation in biological fluids and generally demand tedious post-selection modifications that might compromise binding. One possible solution to warrant biostability is to directly evolve chemically modified aptamers from xenobiotic nucleic acids (XNAs). We have isolated fully modified 2-O-methyl-ribose-1,5-anhydrohexitol nucleic acid (MeORNA-HNA) aptamers targeting the rat vascular endothelial growth factor 164 (rVEGF(164)). Three sequences have been identified that interact with the target protein with affinities in the low-nanomolar range and HNA modifications appeared to be mandatory for their tight binding. The evolution of these XNA aptamers was accomplished using an in vitro selection procedure starting from a fully sugar-modified library containing a 20mer 2-OMe-ribonucleotide region followed by a 47mer HNA sequence. The high binding affinity and selectivity of the selected aptamers were confirmed by several methods including gel-shift, fluorescence polarisation, and enzyme-linked oligonucleotide assays. The isolated HNA ligands exhibited higher specificity to the rVEGF(164) and human VEGF(165) isoforms compared to rat VEGF(120), while very low binding efficiencies were observed to streptavidin and thrombin. Furthermore, it was clearly demonstrated that the resulting aptamers possessed a superior stability to degradation in human serum and DNase I solutions.
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