NDLI: Probing the Mechanisms of DEAD-box Proteins as General RNA Chaperones: The C-terminal Domain of CYT-19 Mediates General Recognition of RNA†

Content Provider	PubMed Central
Author	Grohman, Jacob K. Campo, Mark Del Bhaskaran, Hari Tijerina, Pilar Lambowitz, Alan M. Russell, Rick
Abstract	The DEAD-box protein CYT-19 functions in folding of several group I introns in vivo and a diverse set of group I and group II RNAs in vitro. Recent work using the Tetrahymena group I ribozyme demonstrated that CYT-19 possesses a second RNA binding site, distinct from the unwinding active site, which enhances unwinding activity by binding non-specifically to adjacent RNA structure. Here we probe the region of CYT-19 responsible for that binding by constructing a C-terminal truncation variant that lacks 49 amino acids and terminates at a domain boundary, as defined by limited proteolysis. This truncated protein unwinds a six-base-pair duplex, formed between the oligonucleotide substrate of the Tetrahymena ribozyme and an oligonucleotide corresponding to the internal guide sequence of the ribozyme, with near-wild-type efficiency. However, the truncated protein is activated much less than the wild-type protein when the duplex is covalently linked to the ribozyme or to single-stranded or double-stranded extensions. Thus, the active site for RNA unwinding remains functional in the truncated CYT-19, but the site that binds adjacent RNA structure has been compromised. Equilibrium binding experiments confirmed that the truncated protein binds RNA less tightly than the wild-type protein. RNA binding by the compromised site is important for chaperone activity, as the truncated protein is less active in facilitating folding of a group I intron that requires CYT-19 in vivo. The deleted region contains arginine-rich sequences, as found in other RNA-binding proteins, and may function by tethering CYT-19 to structured RNAs so that it can efficiently disrupt exposed, non-native structural elements, allowing them to re-fold. Many other DExD/H-box proteins also contain arginine-rich ancillary domains, and some of them may function similarly as non-specific RNA-binding elements that enhance general RNA chaperone activity.
Related Links	http://dx.doi.org/10.1021/bi0619472
Ending Page	3022
Page Count	10
Starting Page	3013
File Format	PDF
ISSN	00062960
e-ISSN	15204995
Journal	Biochemistry
Issue Number	11
Volume Number	46
Language	English
Publisher Date	2007-03-01
Access Restriction	Open
Subject Keyword	Biochemistry Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Biochemistry

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Probing the Mechanisms of DEAD-box Proteins as General RNA Chaperones: The C-terminal Domain of CYT-19 Mediates General Recognition of RNA†

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