DEAD‐box proteins as RNA helicases and chaperones

Abstract

DEAD-box proteins are ubiquitous in RNA-mediated processes and function by coupling cycles of ATP binding and hydrolysis to changes in affinity for single-stranded RNA. Many DEAD-box proteins use this basic mechanism as the foundation for a version of RNA helicase activity, efficiently separating the strands of short RNA duplexes in a process that involves little or no translocation. This activity, coupled with mechanisms to direct different DEAD-box proteins to their physiological substrates, allows them to promote RNA folding steps and rearrangements and to accelerate remodeling of RNA-protein complexes. This review will describe the properties of DEAD-box proteins as RNA helicases and the current understanding of how the energy from ATPase activity is used to drive the separation of RNA duplex strands. It will then describe how the basic biochemical properties allow some DEAD-box proteins to function as chaperones by promoting RNA folding reactions, with a focus on the self-splicing group I and group II intron RNAs. KeywordsATPase; CYT-19; DExD/H-box protein; Group I intron; Group II intron; Mss116; RNA folding DEAD-box proteins are the largest family of superfamily 2 (SF2) helicases. They share with other SF2 proteins a helicase core composed of two RecA-like domains and a common set of sequence motifs. In the DEAD-box proteins, motif II includes the sequence D-E-A-D, which is the origin of their name (1). DEAD-box proteins are found in all three domains of life and perform a diverse set of functions in an even more diverse set of cellular processes, including assembly and function of macromolecular machines like the ribosome and spliceosome, quality control mechanisms in gene expression, nuclear export of mRNA, folding of self-splicing RNA introns, and others (2,3) ( Table 1). The broader term "DExD/ H-box" has been used to include other proteins with related motif II sequences (2,3), but a recent comparative sequence analysis has led to questions as to the relevance of this designation, because DEAD-box proteins are not more closely related to DExH helicases than to other families of SF2 helicases (4).In keeping with the broad range of functions for DEAD-box proteins, there is also considerable diversity in their mechanisms. Nevertheless, common themes are emerging. Since their discovery more than 20 years ago (1), the central defining biochemical activity of DEAD-box proteins has been their ability to bind and hydrolyze ATP in a cycle that is stimulated by binding to single-stranded or double-stranded RNA (ssRNA or dsRNA). NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptSubsequent work has shown that the energetic communication implied by the stimulation of ATPase activity by RNA binding forms a common set of properties that underlies the diverse mechanisms of DEAD-box proteins. Progression through the ATPase cycle leads to changes in ssRNA affinity, allowing DEAD-box proteins to bind and release RNA in a regulated manner. This regulated binding is then used by different ...

From unwinding to clamping — the DEAD box RNA helicase family

“…DEAD box proteins are bona fide heli cases that use ATP to bind and unwind RNA duplexes 27 . However, appreciable unwinding is restricted to duplexes with fewer than two helical turns, and many DEAD box helicases unwind only duplexes with fewer than 10 or 12 base pairs 6 . Unwinding efficiency also decreases with increasing duplex stability 28,29 .…”

“…Unwinding efficiency also decreases with increasing duplex stability 28,29 . Singlestranded extensions to duplexes, at either the 3′ or 5′ end, stimulate unwindin g by many DEAD box proteins 1,6,30,31 but in a few instances extensions are not required -at least in vitro -for optima l activity 31 .…”

“…It has become clear that this is because DEAD box proteins do not unwind duplexes based on translocation on the RNA 1,6,28,30,31,33 . Instead, DEAD box proteins load directly onto the duplex region and then pry the strands apart in an ATP-dependent fashion 30,34 .…”

“…For some DEAD box proteins, loading seems to involve multiple protomers of these proteins 34 . Loading of certain DEAD box proteins (for example, CYT19 from Neurospora crassa and bacterial DbpA) can also be facilitate d by structured RNA 6,33 .…”

“…Mutations and deregulation of several DEAD box proteins have been linked to disease states, including cancer 7 . Although all DEAD box proteins contain a structurally highly conserved core with conserved ATP-binding and RNA-binding sites, different proteins have been associated with diverse and seemingly unrelated functions, including the disassembly of RNPs, chaperoning during RNA folding and even stabil ization of protein complexes on RNA 5,6 . How these highly conserved proteins fulfil such an array of different functions has been a long-standing question.…”