1. Potent 2′-amino-, and 2′-fluoro-2′- deoxyribonucleotide RNA inhibitors of keratinocyte growth factor
- Author
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Susan Jennings, Ying-Fon Chang, Tim Fitzwater, Carol Bell, Derek Jellinek, Nikos Pagratis, and Chinh Dang
- Subjects
Fibroblast Growth Factor 7 ,Deoxyribonucleotides ,Molecular Sequence Data ,Biomedical Engineering ,Bioengineering ,Applied Microbiology and Biotechnology ,Substrate Specificity ,Mice ,Deoxyribonucleotide ,chemistry.chemical_compound ,Ribonucleases ,Drug Stability ,Animals ,Humans ,Binding site ,Growth Substances ,Nuclease ,Binding Sites ,Base Sequence ,biology ,Oligonucleotide ,Temperature ,RNA ,3T3 Cells ,Affinities ,Fibroblast Growth Factors ,Biochemistry ,chemistry ,biology.protein ,Nucleic Acid Conformation ,Molecular Medicine ,Pancreatic ribonuclease ,Fibroblast Growth Factor 10 ,Systematic evolution of ligands by exponential enrichment ,Biotechnology - Abstract
Reiterative in vitro selection-amplification from random oligonucleotide libraries allows the identification of molecules with specific functions such as binding to specific proteins. The therapeutic usefulness of such molecules depends on their high affinity and nuclease resistance. Libraries of RNA molecules containing 2'amino-(2'NH2)- or 2'fluoro-(2'F)-2'-deoxypyrimidines could yield ligands with similar nuclease resistance but not necessarily with similar affinities. This is because the intramolecular helices containing 2'NH2 have lower melting temperatures (Tm) compared with helices containing 2'F, giving them thermodynamically less stable structures and possibly weaker affinities. We tested these ideas by isolating high-affinity ligands to human keratinocyte growth factor from libraries containing modified RNA molecules with either 2'NH2 or 2'F pyrimidines. We demonstrated that 2'F RNA ligands have affinities (Kd approximately 0.3-3 pM) and bioactivities (Ki approximately 34 pM) superior to 2'NH2 ligands (Kd approximately 400 pM and Ki approximately 10 nM). In addition, 2'F ligands have extreme thermo-stabilities (Tm approximately 78 degrees C in low salt, and specificities).
- Published
- 1997