Berlin 2005 – scientific programme

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AKB: Biologische Physik

AKB 100: Poster Session I

AKB 100.4: Poster

Saturday, March 5, 2005, 16:45–18:45, Poster TU D

Single Molecule FRET Experiments with the RNA Helicase YxiN — •Bettina Theißen and Dagmar Klostermeier — Experimentalphysik IV, Universität Bayreuth

RNA helicases unwind double helical RNA structures in an ATP-dependant manner. Their function is essential for all cellular processes that require structural reorganisation of RNA, for example transcription, translation or ribosome biogenesis. While little is known about the mechanism of unwinding, an opening and closing movement of a cleft between two domains during the catalytic cycle has been proposed. Such a movement leads to changes of intramolecular distances. Since fluorescence resonance energy transfer (FRET) can be used to measure distances between two fluorophores, it is a suitable tool for studying these conformational changes.

YxiN is a RNA-helicase from Bacillus subtilis that is involved in ribosome biogenesis. For FRET experiments the donor fluorophore alexa 488 and the acceptor fluorophore tetramethylrhodamine have been coupled site-specifically to genetically engineered cysteines on both sides of the cleft. With this doubly labelled protein we perform single molecule FRET experiments to directly observe conformational changes during unwinding of the RNA. These will lead to a detailed understanding of the catalytic mechanism of RNA-helicases.