Single-molecule kinetics of the eukaryotic initiation factor 4AI upon RNA unwinding
Details
Publication Year 2014-07-08, Volume 22, Issue #7, Page 941-8
Journal Title
Structure
Publication Type
Journal Article
Abstract
The eukaryotic translation initiation factor 4AI (eIF4AI) is the prototypical DEAD-box RNA helicase. It has a "dumbbell" structure consisting of two domains connected by a flexible linker. Previous studies demonstrated that eIF4AI, in conjunction with eIF4H, bind to loop structures and repetitively unwind RNA hairpins. Here, we probe the conformational dynamics of eIF4AI in real time using single-molecule FRET. We demonstrate that eIF4AI/eIF4H complex can repetitively unwind RNA hairpins by transitioning between an eIF4AI "open" and a "closed" conformation using the energy derived from ATP hydrolysis. Our experiments directly track the conformational changes in the catalytic cycle of eIF4AI and eIF4H, and this correlates precisely with the kinetics of RNA unwinding. Furthermore, we show that the small-molecule eIF4A inhibitor hippuristanol locks eIF4AI in the closed conformation, thus efficiently inhibiting RNA unwinding. These results indicate that the large conformational changes undertaken by eIF4A during the helicase catalytic cycle are rate limiting.
Publisher
Cell Press
WEHI Research Division(s)
Cell Signalling And Cell Death
Rights Notice
Refer to copyright notice on published article.


Creation Date: 2015-03-12 03:59:02
Last Modified: 2015-03-16 09:21:14
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