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Isolation of high-affinity RNA ligands to HIV-1 integrase from a random pool
Isolation of high-affinity RNA ligands to HIV-1 integrase from a random pool.
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We were able to isolate high-affinity RNAs from a random pool that binds to integrase protein from the human immunodeficiency virus-type 1 using the procedure now known as SELEX. Generally, the RNAs fell into three different classes in binding buffer containing 250 mM NaCl: group I class of molecules binds integrase with a dissociation constant (Kd) on the order of 10 nM, group II molecules had a Kd of about 80 nM, and group III about 800 nM. The RNA with the highest affinity from the group I class of molecules, designated P5, was characterized using computer modeling, chemical and enzymatic probing, and deletion analysis. Our secondary structure model for this RNA suggests interactions between looped-out fixed nucleotides and nucleotides from the randomized region; a GNRA tetraloop is also in the structure. We showed that our integrase was able to process a U5 mimic in vitro. P5 competes effectively for binding with the double-stranded DNA mimic of U5 at 180 mM NaCl concentration.
Allen P, Worland S, Gold L
Virology
1995-06-01 00:00
209
2
327-36
Base Sequence,Cloning, Molecular,Computer Simulation,DNA Nucleotidyltransferases,DNA Primers,Genes, Viral,HIV-1,Integrases,Kinetics,Ligands,Models, Molecular,Molecular Sequence Data,Nucleic Acid Conformation,Polymerase Chain Reaction,Protein Binding,RNA, Viral,Recombinant Proteins,Substrate Specificity,Viral Structural Proteins,Virus Integration,DNA Primers,Ligands,RNA, Viral,Recombinant Proteins,Viral Structural Proteins,DNA Nucleotidyltransferases,Integrases
Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder 80309-0347, USA
Virology
NIGMS GM19963, NIGMS GM28685, NIGMS GM42651
0042-6822
10.1006/viro.1995.1264
S0042-6822(85)71264-0
417
True
7778267
