The discovery of RNA sequences of potential biomedical importance has dramatically outpaced chemists'ability to design and synthesize novel selective RNA-binding compounds. This is due largely to a gap in knowledge in the field with regard to fundamental determinants of selectivity. This proposal seeks to test the hypothesis that the sequence selectivity of an RNA-binding compound is directly related to its kinetic off rate or """"""""residence time"""""""" in the desired binding site. While a generally accepted principle in the realm of protein and enzyme recognition, and tested also in the context of DNA recognition, to our knowledge this concept has not been applied to compounds binding RNA. This hypothesis will be tested via three Aims. First, well-validated (but low- throughput) techniques will be used to analyze the binding properties of a series of known RNA-targeted compounds. Second, a new analytical methodology developed in our laboratory termed Arrayed Imaging Reflectometry will be tested in the context of multiplex (high-throughput) assessment of RNA-binding kinetic constants. This will also involve the development of new statistical methods for the analysis of time-dependent array data. Third, we will examine the effect of systematic functional group modification on the binding kinetics and sequence selectivity of a novel compound discovered in our lab that targets a viral RNA critical to the HIV life cycle. Completion of the proposed research will provide a new paradigm for RNA-targeted molecular design based on consideration of binding kinetics, as well as a new analytical tool for high-throughput characterization of RNA binding, and new lead compounds targeting HIV.
RNA is rapidly emerging as an important target for drug development. However, in contrast to proteins and DNA, our understanding of the factors that lead to the successful design of an RNA-targeted small molecule are still at an early stage. In the proposed research, we will develop new tools that will dramatically increase the speed of analyzing small molecule - RNA interactions, while testing a novel hypothesis regarding RNA-binding selectivity. These methods will be applied in the context of new molecules targeting HIV.
|Hilimire, Thomas A; Bennett, Ryan P; Stewart, Ryan A et al. (2016) N-Methylation as a Strategy for Enhancing the Affinity and Selectivity of RNA-binding Peptides: Application to the HIV-1 Frameshift-Stimulating RNA. ACS Chem Biol 11:88-94|
|McAnany, John D; Reichert, John P; Miller, Benjamin L (2016) Probing the geometric constraints of RNA binding via dynamic covalent chemistry. Bioorg Med Chem 24:3940-6|
|Lifson, Mark A; Carter, Jared A; Miller, Benjamin L (2015) Functionalized Polymer Microgel Particles Enable Customizable Production of Label-Free Sensor Arrays. Anal Chem 87:7887-93|
|Liu, Zhi-Ping; Wu, Canglin; Miao, Hongyu et al. (2015) RegNetwork: an integrated database of transcriptional and post-transcriptional regulatory networks in human and mouse. Database (Oxford) 2015:|
|Yadav, Amrita R; Mace, Charles R; Miller, Benjamin L (2014) Examining the interactions of the splicing factor MBNL1 with target RNA sequences via a label-free, multiplex method. Anal Chem 86:1067-75|
|Yadav, Amrita R; Sriram, Rashmi; Carter, Jared A et al. (2014) Comparative study of solution-phase and vapor-phase deposition of aminosilanes on silicon dioxide surfaces. Mater Sci Eng C Mater Biol Appl 35:283-90|
|Liu, Zhi-Ping; Wu, Hulin; Zhu, Jian et al. (2014) Systematic identification of transcriptional and post-transcriptional regulations in human respiratory epithelial cells during influenza A virus infection. BMC Bioinformatics 15:336|
|Hoskins, Jason W; Ofori, Leslie O; Chen, Catherine Z et al. (2014) Lomofungin and dilomofungin: inhibitors of MBNL1-CUG RNA binding with distinct cellular effects. Nucleic Acids Res 42:6591-602|
|Ofori, Leslie O; Hilimire, Thomas A; Bennett, Ryan P et al. (2014) High-affinity recognition of HIV-1 frameshift-stimulating RNA alters frameshifting in vitro and interferes with HIV-1 infectivity. J Med Chem 57:723-32|