eIF4G stimulates the activity of the DEAD box protein eIF4A by a conformational guidance mechanism

Hilbert, Manuel; Kebbel, Fabian; Gubaev, Airat; Klostermeier, Dagmar

doi:10.1093/nar/gkq1127

Cited by 103 publications

(153 citation statements)

References 39 publications

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“…However, the two core domains do not close completely, as observed for eIF4AIII in the EJC 89,91 . The eIF4G-induced, half-open form of eIF4A promotes RNA binding and also accelerates phosphate release, which is presumably the rate-limiting step in the ATPase cycle for eIF4A 91 . These findings are consistent with the well-documented stimulation of the RNA-dependent ATPase activity of eIF4A by eIF4G 85 .…”

Section: Nuclear Specklesmentioning

confidence: 82%

From unwinding to clamping — the DEAD box RNA helicase family

Linder

Jankowsky

2011

Nat Rev Mol Cell Biol

988

1,069

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Nearly all aspects of RNA metabolism, from transcription and translation to mRNA decay, involve RNA helicases, which are enzymes that use ATP to bind or remodel RNA and RNA-protein complexes (ribonucleoprotein (RNP) complexes) 1 . RNA helicases are found in all three domains of life, and many viruses also encode one or more of these proteins 2,3 . Together with the structurally related DNA helicases that function in replication, recombination and repair, the RNA helicases are classified into superfamilies and families, based on sequence and structural features 3,4 . DEAD box proteins form the largest helicase family, with 37 members in humans and 26 in Saccharomyces cerevisiae 3 , and are characterized by the presence of an Asp-Glu-Ala-Asp (DEAD) motif.DEAD box helicases have central and, in many cases, essential physiological roles in cellular RNA metabolism 5 (FIG. 1). The proteins generally function as part of larger multicomponent assemblies, such as the spliceosom e or the eukaryotic translation initiation machinery 6 . 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.Research has now started to illuminate the molecular basis for this functional diversity. In this Review, we summarize how structural data, together with biochemical and biophysical studies, have revealed unexpected modes by which these proteins function. We also discuss how novel molecular and cell biological approaches have better defined the cellular roles of DEAD box helicases. We outline our current view on common structural and mechan istic themes that have emerged, and on physical models of how these 'unusual' RNA helicases work.Structures of DEAD box RNA helicases A number of structural studies have universally shown that members of the DEAD box family contain a highly conserved helicase core that harbours the binding sites for ATP and RNA 3,4,8 . The core is surrounded by variable auxiliary domains, which are thought to be critical for the diverse functions of these enzymes.Structure of the helicase core. DEAD box proteins belong to helicase superfamily 2 (SF2) 2 . Similarly to all SF2 helicases, DEAD box proteins are built around a highly conserved helicase core of two virtually identical domains that resemble the bacterial recombination protein recombinase A (RecA) 3,4,8 (FIG. 2a,b). Within this helicase core, at least 12 characteristic sequence motifs are located at conserved positions (FIG. 2a,b). Some of these motifs are conserved across the entire SF2 family, whereas others are found only in the DEAD box family 3 ....

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Section: Nuclear Specklesmentioning

confidence: 82%

From unwinding to clamping — the DEAD box RNA helicase family

Linder

Jankowsky

2011

Nat Rev Mol Cell Biol

988

1,069

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“…Interestingly, cofactors are often RNA-binding proteins (19)(20)(21)(22)(23)(24)(25). We and others therefore postulated that these cofactors could also function to provide specificity for DEAD-box and DEAH-box proteins (1, 18).Ded1 and eIF4A, two of the best-studied DEAD-box proteins (19,24,(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38), have known binding cofactors: Gle1 and eIF4G, respectively. Because Ded1 and eIF4A both function in translation initiation, a process that requires the recognition of many diverse mRNAs, sequence specificity for these particular DEADbox proteins may be undesirable.…”

mentioning

confidence: 99%

Cofactor-dependent specificity of a DEAD-box protein

Young

Khoshnevis

Karbstein

2013

Proc. Natl. Acad. Sci. U.S.A.

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DEAD-box proteins, a large class of RNA-dependent ATPases, regulate all aspects of gene expression and RNA metabolism. They can facilitate dissociation of RNA duplexes and remodeling of RNA-protein complexes, serve as ATP-dependent RNA-binding proteins, or even anneal duplexes. These proteins have highly conserved sequence elements that are contained within two RecA-like domains; consequently, their structures are nearly identical. Furthermore, crystal structures of DEAD-box proteins with bound RNA reveal interactions exclusively between the protein and the RNA backbone. Together, these findings suggest that DEAD-box proteins interact with their substrates in a nonspecific manner, which is confirmed in biochemical experiments. Nevertheless, this contrasts with the need to target these enzymes to specific substrates in vivo. Using the DEAD-box protein Rok1 and its cofactor Rrp5, which both function during maturation of the small ribosomal subunit, we show here that Rrp5 provides specificity to the otherwise nonspecific biochemical activities of the Rok1 DEAD-domain. This finding could reconcile the need for specific substrate binding of some DEAD-box proteins with their nonspecific binding surface and expands the potential roles of cofactors to specificity factors. Identification of helicase cofactors and their RNA substrates could therefore help define the undescribed roles of the 19 DEAD-box proteins that function in ribosome assembly.RNA helicases | annealing D EAD-box proteins are RNA-binding ATPases that are involved in all aspects of RNA metabolism: translation initiation, pre-mRNA splicing, mRNA export and decay, and ribosome biogenesis. In these processes, their functions include RNA duplex unwinding, RNA-protein complex remodeling, RNA duplex annealing, and ATP-dependent RNA binding (1-5).In vivo, these enzymes have specific functions that often involve the recognition of specific RNAs and the discrimination against a myriad of nonsubstrates. Nevertheless, only DbpA, a DEAD-box protein involved in bacterial ribosome assembly (6-8), has sequence-specific ATPase activity (6, 9), which arises from unique sequences in its C terminus (10). Such sequence specificity has not been demonstrated for any other DEAD-box protein and is consistent with their highly conserved structures and the almost universal conservation of residues that contact the RNA. Furthermore, analysis of the structures of RNA-bound DEAD-box proteins reveals that RNA contacts are made with the sugarphosphate backbone (11-17), largely precluding sequence-specific interactions between DEAD-box proteins and RNA.Many DEAD-box and related DEAH-box proteins function in conjunction with cofactors (ref. 18 and references therein). These cofactors can regulate the activity of DEAD-box or DEAH-box proteins by stimulating or inhibiting their ATPase, helicase, RNAbinding, or nucleotide-binding activities. Interestingly, cofactors are often RNA-binding proteins (19)(20)(21)(22)(23)(24)(25). We and others therefore postulated that these cofactors could als...

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“…We have previously validated our correction procedure and have shown that it yields corrected FRET efficiencies that reflect correct intermolecular distances (see Ref. 24 for a detailed description).…”

Section: Methodsmentioning

confidence: 99%

Potassium Ions Are Required for Nucleotide-induced Closure of Gyrase N-gate

Gubaev

Klostermeier

2012

Journal of Biological Chemistry

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Background: ATP-dependent closure of the N-gate is a key step for negative DNA supercoiling by gyrase. Results: Potassium ions are required for efficient nucleotide-induced N-gate closure and DNA supercoiling but are dispensable for DNA relaxation. Conclusion: Potassium ions help coordinate the nucleotide cycle to gate movements of gyrase. Significance: Coordinated conformational changes are crucial for the function of ATP-driven molecular machines in general.DNA gyrase catalyzes ATP-dependent negative supercoiling of DNA by a strand passage mechanism that requires coordinated opening and closing of three protein interfaces, the N-, DNA-, and C-gates. ATP binding to the GyrB subunits of gyrase causes dimerization and N-gate closure. The closure of the N-gate is a key step in the gyrase catalytic cycle, as it captures the DNA segment to be transported and poises gyrase toward strand passage. We show here that K ؉ ions are required for DNA supercoiling but are dispensable for ATP-independent DNA relaxation. Although DNA binding, distortion, wrapping, and DNA-induced narrowing of the N-gate occur in the absence of K ؉ , nucleotide-induced N-gate closure depends on their presence. Our results provide evidence that K ؉ ions relay small conformational changes in the nucleotide-binding pocket to the formation of a tight dimer interface at the N-gate by connecting regions from both GyrB monomers and suggest an important role for K ؉ in synchronization of N-gate closure and DNA-gate opening.Gyrase is a DNA topoisomerase that introduces negative supercoils into plasmid DNA in an ATP-dependent reaction (1) and plays an important role in DNA replication, transcription, and recombination (2). The active unit of gyrase is composed of two GyrA and two GyrB subunits that assemble into an A 2 B 2 heterotetramer. Gyrase forms two cavities, delimited by three protein interfaces termed the N-, DNA-, and C-gates (see Fig. 1A). During ATP-dependent negative supercoiling, these gates open and close in a coordinated manner to allow for strand passage toward negative DNA supercoiling (3-8). The gyrase catalytic cycle begins when a gate DNA (G-segment) is bound at the DNA-gate and distorted (7). Interaction of DNA flanking the G-segment with the C-terminal domains (CTDs) 2 causes the CTDs to move upward and sideways (9), and complete wrapping of DNA leads to narrowing of the N-gate formed by the GyrB subunits (8). The N-gate acts as an ATPdependent clamp (10): ATP binding to the ATPase domains of the GyrB subunits induces GyrB dimerization and N-gate closure (8,(11)(12)(13), leading to the trapping of the transport DNA (T-segment). N-gate closure and DNA-gate opening appear to be coupled, with possible contributions from the T-segment (10). After passage of the T-segment through the gap in the G-segment, the G-segment is religated, the T-segment leaves the enzyme through the C-gate formed by GyrA (14,15), and the N-gate reopens (8, 12). Our recent results provide evidence for a bidirectional communication between the N-and DNAgates of g...

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eIF4G stimulates the activity of the DEAD box protein eIF4A by a conformational guidance mechanism

Cited by 103 publications

References 39 publications

From unwinding to clamping — the DEAD box RNA helicase family

From unwinding to clamping — the DEAD box RNA helicase family

Cofactor-dependent specificity of a DEAD-box protein

Potassium Ions Are Required for Nucleotide-induced Closure of Gyrase N-gate

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