Andrei Yu Kobitski

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Using a combination of advanced RNA synthesis techniques and single molecule spectroscopy, the deconvolution of individual contributions of posttranscriptional modifications to the overall folding and stabilization of human mitochondrial tRNA(Lys) is described. An unexpected destabilizing effect of two pseudouridines on the native tRNA folding was(More)
Investigation of single RNA molecules using fluorescence resonance energy transfer (FRET) is a powerful approach to investigate dynamic and thermodynamic aspects of the folding process of a given RNA. Its application requires interdisciplinary work from the fields of chemistry, biochemistry, and physics. The present work gives detailed instructions on the(More)
Enzymology at the single-molecule level by using fluorescence resonance energy transfer (smFRET) offers unprecedented insight into mechanistic aspects of catalytic reactions. Implementing spatiotemporal control of the reaction by using an external trigger is highly valuable in these challenging experiments. Here, we have incorporated a light-cleavable(More)
Single-molecule fluorescence microscopy experiments on RNA molecules brought to light the highly complex dynamics of key biological processes, including RNA folding, catalysis of ribozymes, ligand sensing of riboswitches and aptamers, and protein synthesis in the ribosome. By using highly advanced biophysical spectroscopy techniques in combination with(More)
The Diels-Alder reaction is one of the most important C-C bond-forming reactions in organic chemistry, and much effort has been devoted to controlling its enantio- and diastereoselectivity. The Diels-Alderase ribozyme (DAse) catalyses the reaction between anthracene dienes and maleimide dienophiles with multiple-turnover, stereoselectivity, and up to(More)
The folding energy landscape of RNA is greatly affected by interactions between the RNA and counterions that neutralize the backbone negative charges and may also participate in tertiary contacts. Valence, size, coordination number, and electron shell structure can all contribute to the energetic stabilization of specific RNA conformations. Using(More)
Fluorescence resonance energy transfer (FRET) is a superb technique for measuring conformational changes of proteins on the single molecule level (smFRET) in real time. It requires introducing a donor and acceptor fluorophore pair at specific locations on the protein molecule of interest, which has often been a challenging task. By using two different(More)
We have studied the folding kinetics of the core intermediate (I) state of RNase H by using a combination of single-molecule FRET (smFRET) and hidden Markov model analysis. To measure fast dynamics in thermal equilibrium as a function of the concentration of the denaturant GdmCl, a special FRET labeled variant, RNase H 60-113, which is sensitive to folding(More)