期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 105, 期 51, 页码 20203-20208出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0811075106
关键词
CYT-19; group I intron; RNA chaperone; RNA folding; RNA helicase
资金
- National Institutes of Health [GM070456, GM037951]
- Welch Foundation [F-1563]
- National Institutes of Health Postdoctoral Fellowship [F32-GM76961]
DEAD-box proteins are ubiquitous in RNA metabolism and use ATP to mediate RNA conformational changes. These proteins have been suggested to use a fundamentally different mechanism from the related DNA and RNA helicases, generating local strand separation while remaining tethered through additional interactions with structured RNAs and RNA-protein (RNP) complexes. Here, we provide a critical test of this model by measuring the number of ATP molecules hydrolyzed by DEAD-box proteins as they separate short RNA helices characteristic of structured RNAs (6-11 bp). We show that the DEAD-box protein CYT-19 can achieve complete strand separation using a single ATP, and that 2 related proteins, Mss116p and Ded1p, display similar behavior. Under some conditions, considerably < 1 ATP is hydrolyzed per separation event, even though strand separation is strongly dependent on ATP and is not supported by the nucleotide analog AMP-PNP. Thus, ATP strongly enhances strand separation activity even without being hydrolyzed, most likely by eliciting or stabilizing a protein conformation that promotes strand separation, and AMP-PNP does not mimic ATP in this regard. Together, our results show that DEAD-box proteins can disrupt short duplexes by using a single cycle of ATP-dependent conformational changes, strongly supporting and extending models in which DEAD-box proteins perform local rearrangements while remaining tethered to their target RNAs or RNP complexes. This mechanism may underlie the functions of DEAD-box proteins by allowing them to generate local rearrangements without disrupting the global structures of their targets.
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