The goals of this proposed work are to develop and optimize the properties of a promising new binding motif for single-stranded polynucleotides, and to utilize this information to construct molecules which may have anti- retroviral properties. The circular ligands described in this proposal are designed to block gene expression for specific sequences of viral RNA by binding exceptionally tightly to these sequences. It has been shown in published reports that linear oligonucleotides and their analogs possess some of these inhibitory properties. Circular oligonucleotides or their analogs may prove to have sufficient advantages over other oligonucleotides that they will eventually be useful as drugs against viral infection. These long-term goals will require that binding by these ligands be studied on a molecular basis before they can be tested in vitro and in vivo for inhibitory activity against expression of HIV-1 genes. Experiments planned for the term spanned by this proposal include both molecular and structural studies as well as applications of the most promising ligands. First, optimization of the yield of the reaction which converts linear precursors into circles will be carried out. Second, structural optimization of the ligands, including size and sequence and their effects on binding strength will be studied. Also examined will be the extent of target sequence variability in RNA strands, as well as the level of sequence specificity that can be achieved in binding. Studies of the nuclease susceptibility of oligonucleotides consisting of natural nucleotides and of synthetically modified ones will add important practical information. Finally, the ligands will be examined for their ability to inhibit transcription and translation of specific genetic sequences in vitro. The insight gained from these proposed experiments should allow the design of optimized molecules which have the potential to act as inhibitors of retroviral replication.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM046625-03
Application #
3306059
Study Section
AIDS and Related Research Study Section 4 (ARRD)
Project Start
1991-07-01
Project End
1994-06-30
Budget Start
1993-07-01
Budget End
1994-06-30
Support Year
3
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of Rochester
Department
Type
Schools of Arts and Sciences
DUNS #
208469486
City
Rochester
State
NY
Country
United States
Zip Code
14627
Mohsen, Michael G; Kool, Eric T (2016) The Discovery of Rolling Circle Amplification and Rolling Circle Transcription. Acc Chem Res 49:2540-2550
Ohmichi, T; Kool, E T (2000) The virtues of self-binding: high sequence specificity for RNA cleavage by self-processed hammerhead ribozymes. Nucleic Acids Res 28:776-83
Xu, Y; Kool, E T (1999) High sequence fidelity in a non-enzymatic DNA autoligation reaction. Nucleic Acids Res 27:875-81
Xu, Y; Kool, E T (1998) Chemical and enzymatic properties of bridging 5'-S-phosphorothioester linkages in DNA. Nucleic Acids Res 26:3159-64
Wang, S; Xu, Y; Kool, E T (1997) Recognition of RNA by triplex formation: divergent effects of pyrimidine C-5 methylation. Bioorg Med Chem 5:1043-50
Kool, E T (1996) Circular oligonucleotides: new concepts in oligonucleotide design. Annu Rev Biophys Biomol Struct 25:1-28
Wang, S; Friedman, A E; Kool, E T (1995) Origins of high sequence selectivity: a stopped-flow kinetics study of DNA/RNA hybridization by duplex- and triplex-forming oligonucleotides. Biochemistry 34:9774-84
Wang, S; Kool, E T (1994) Circular RNA oligonucleotides. Synthesis, nucleic acid binding properties, and a comparison with circular DNAs. Nucleic Acids Res 22:2326-33
Booher, M A; Wang, S; Kool, E T (1994) Base pairing and steric interactions between pyrimidine strand bridging loops and the purine strand in DNA pyrimidine.purine.pyrimidine triplexes. Biochemistry 33:4645-51
Wang, S; Booher, M A; Kool, E T (1994) Stabilities of nucleotide loops bridging the pyrimidine strands in DNA pyrimidine.purine.pyrimidine triplexes: special stability of the CTTTG loop. Biochemistry 33:4639-44

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