Whenever an important molecular tool is developed, new discoveries follow. Fluorescent and luminescent proteins are prime examples of tools that have had an enormous impact on biology. However, molecular biology and biophysics lack similar molecules that allow direct interrogation of the great variety of cellular functional RNAs. This project aims to develop fluorogenic aptamers and ribozymes, RNAs that can be detected with high spatial and temporal resolution in live cells, as well as in vitro. These clonable modules will also find applications in structural biology, particularly in NMR, X-ray crystallography, and luminescence-based measurements. Our goal is to use in vitro selections to identify RNA modules that bind chelated lanthanides with high affinity and selectivity. In addition, we will develop new method for in vitro selection of multiple-turnover enzymes and use this method to identify fluorogenic multiple-turnover ribozymes. The technique will then be used to discover new fluorophores together with the enzymes that catalyze their synthesis and will thus significantly extend the range of experiments that in vitro selections can be applied to. All together, this project will significantly improve our ability to analyze RNAs in vitro and in vivo, and will create methods for de novo selection of highly efficient multiple-turnover enzymes.
Ribonucleic acids are involved in many fundamental processes in biological systems. This project will develop new tools for efficient analysis of RNAs in live tissues and provide a platform the analysis of fundamental biological processes involving cellular and viral RNAs.
|Burke, Cassandra R; Lupták, Andrej (2018) DNA synthesis from diphosphate substrates by DNA polymerases. Proc Natl Acad Sci U S A 115:980-985|
|Webb, Chiu-Ho T; Lupták, Andrej (2018) Kinetic Parameters of trans Scission by Extended HDV-like Ribozymes and the Prospect for the Discovery of Genomic trans-Cleaving RNAs. Biochemistry 57:1440-1450|
|Pobanz, Kelsey; Lupták, Andrej (2016) Improving the odds: Influence of starting pools on in vitro selection outcomes. Methods 106:14-20|
|Rampášek, Ladislav; Jimenez, Randi M; Lupták, Andrej et al. (2016) RNA motif search with data-driven element ordering. BMC Bioinformatics 17:216|
|Long, Anthony; Liti, Gianni; Luptak, Andrej et al. (2015) Elucidating the molecular architecture of adaptation via evolve and resequence experiments. Nat Rev Genet 16:567-82|
|Vu, Michael M K; Jameson, Nora E; Masuda, Stuart J et al. (2012) Convergent evolution of adenosine aptamers spanning bacterial, human, and random sequences revealed by structure-based bioinformatics and genomic SELEX. Chem Biol 19:1247-54|
|Jimenez, Randi M; Rampasek, Ladislav; Brejova, Brona et al. (2012) Discovery of RNA motifs using a computational pipeline that allows insertions in paired regions and filtering of candidate sequences. Methods Mol Biol 848:145-58|
|Ruminski, Dana J; Webb, Chiu-Ho T; Riccitelli, Nathan J et al. (2011) Processing and translation initiation of non-long terminal repeat retrotransposons by hepatitis delta virus (HDV)-like self-cleaving ribozymes. J Biol Chem 286:41286-95|
|Sendroiu, Iuliana E; Gifford, Lida K; Lupták, Andrej et al. (2011) Ultrasensitive DNA microarray biosensing via in situ RNA transcription-based amplification and nanoparticle-enhanced SPR imaging. J Am Chem Soc 133:4271-3|
|Jimenez, Randi M; Delwart, Eric; Lupták, Andrej (2011) Structure-based search reveals hammerhead ribozymes in the human microbiome. J Biol Chem 286:7737-43|
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