Structural insights into the mechanism of the DEAH-box RNA helicase Prp43

Tauchert, M.J.; Fourmann, Jean-Baptiste; Lührmann, Reinhard; Ficner, Ralf

doi:10.7554/elife.21510

Cited by 90 publications

(238 citation statements)

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“…The basic features of this mechanism are the same as determined for other members of the DEAH-box family (31,33). These basic features, including a requirement for translocation, probably reflect the presence in DEAH-box proteins of C-terminal domains that surround nucleic acid substrates and are likely important for translocation (33,(43)(44)(45), as well as conserved sequence motifs within the helicase core. While we established this mechanism using principally DNA G4s, analogous RNA constructs were processed by DHX36 with similar rates, indicating that DHX36 likely uses the same mechanism to disrupt DNA and RNA G4s.…”

Section: Discussionmentioning

confidence: 60%

“…With rate-limiting substrate binding, DHX36 can be described as having achieved a limited definition of 'catalytic perfection' for disrupting these G4s, as any further improvements in the rates of first-order steps would not increase the reaction efficiency or the level of specificity (46). The binding rate constants of ~2 × 10 8 M -1 min -1 for tetramolecular G4s and ~1 × 10 9 M -1 min -1 for unimolecular G4s are 2-3 orders of magnitude below the encounter frequency, most likely owing to the complexity of accommodating the 3′ extension within a crevice of the protein that may be only transiently accessible (45).…”

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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The G-quadruplex (G4) resolvase DHX36 efficiently and specifically disrupts DNA G4s via a translocation-based helicase mechanism

Yangyuoru

Bradburn

et al. 2018

Journal of Biological Chemistry

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Single-stranded DNA (ssDNA) and RNA regions that include at least four closely spaced runs of three or more consecutive guanosines strongly tend to fold into stable G-quadruplexes (G4s). G4s play key roles as DNA regulatory sites and as kinetic traps that can inhibit biological processes, but how G4s are regulated in cells remains largely unknown. Here, we developed a kinetic framework for G4 disruption by DEAH-box helicase 36 (DHX36), the dominant G4 resolvase in human cells. Using tetramolecular DNA and RNA G4s with four to six G-quartets, we found that DHX36-mediated disruption is highly efficient, with rates that depend on G4 length under saturating conditions (k cat ) but not under subsaturating conditions (k cat /K M ). These results suggest that a step during G4 disruption limits the k cat value and that DHX36 binding limits k cat /K M . Similar results were obtained for unimolecular DNA G4s. DHX36 activity depended on a 3′ ssDNA extension and was blocked by a polyethylene glycol linker, indicating that DHX36 loads onto the extension and translocates 3′-5′ toward the G4. DHX36 unwound dsDNA poorly compared with G4s of comparable intrinsic lifetime. Interestingly, we observed that DHX36 has striking 3′-extension sequence preferences that differ for G4 disruption and dsDNA unwinding, most likely arising from differences in the rate-limiting step for the two activities. Our results indicate that DHX36 disrupts G4s with a conventional helicase mechanism that is tuned for great efficiency and specificity for G4s. The dependence of DHX36 on the 3′-extension sequence suggests that the extent of formation of genomic G4s may not track directly with G4 stability. ___________________________________Single-stranded DNA and RNA segments that include at least four closely-spaced runs of three or more consecutive guanosine nucleotides have a strong intrinsic propensity to fold into stable G-quadruplex structures (G4s) (1) (Fig. 1A, top left). The genomes of humans and many other eukaryotes are replete with putative G4-forming sequences, and pronounced, conserved patterns have been observed in their distribution, suggesting that Gquadruplex structures form at least transiently in cells and perform conserved functions (2-6). Supporting this hypothesis, G4s have been detected in cells for both .To function as regulatory elements, G4s must be folded and disrupted in a regulated way. In addition, processes that require single-stranded DNA or RNA, such as replication and translation, require that G4s be temporarily disrupted. The high stability and long intrinsic lifetime of G4s suggest Mechanism of G4 disruption by DHX362 that their efficient disruption requires the activity of ATP-dependent enzymes such as helicases.Supporting this view, incorporation of sequences predicted to form particularly stable G4s leads to genetic instability in yeast, suggesting that these G4 structures are not processed efficiently and pose blocks to replication (12). Indeed, several helicases have been shown to possess G4 disruption acti...

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Section: Discussionmentioning

confidence: 60%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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The G-quadruplex (G4) resolvase DHX36 efficiently and specifically disrupts DNA G4s via a translocation-based helicase mechanism

Yangyuoru

Bradburn

et al. 2018

Journal of Biological Chemistry

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“…Recent structures of Prp43 from C. thermophilum have provided insight into how DEAH-box proteins load onto ssRNA adjacent to their target structures to initiate unwinding (57). Among other roles, Prp43 is required in the final step of pre-mRNA splicing, where it disrupts base pairing between the U2 snRNP and the intron branch point, releasing the U2, U5, and U6 snRNPs from the lariat intron (58–62).…”

Section: Deah-box Proteins As Molecular Winchesmentioning

confidence: 99%

“…Among other roles, Prp43 is required in the final step of pre-mRNA splicing, where it disrupts base pairing between the U2 snRNP and the intron branch point, releasing the U2, U5, and U6 snRNPs from the lariat intron (58–62). Like other DEAH-box proteins, Prp43 lacks sequence specificity for RNA substrates, a property that was rationalized from the crystal structure of the enzyme bound to ADP-BeF 3 and U 7 RNA, as the majority of contacts are formed with the sugar-phosphate backbone of the RNA, not the uracil bases (57, 63). A similar structure without RNA showed that large conformational changes in the C-terminal domains, particularly the ratchet-like and OB-fold domains, lead to an opening of the RNA-binding tunnel between D2 and the C-terminal domains (Fig.…”

Section: Deah-box Proteins As Molecular Winchesmentioning

confidence: 99%

Distinct RNA-unwinding mechanisms of DEAD-box and DEAH-box RNA helicase proteins in remodeling structured RNAs and RNPs

Gilman

Tijerina

Russell

2017

Biochemical Society Transactions

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Structured RNAs and RNA-protein complexes (RNPs) fold through complex pathways that are replete with misfolded traps, and many RNAs and RNPs undergo extensive conformational changes during their functional cycles. These folding steps and conformational transitions are frequently promoted by RNA chaperone proteins, notably by superfamily 2 (SF2) RNA helicase proteins. The two largest families of SF2 helicases, DEAD-box and DEAH-box proteins, share evolutionarily conserved helicase cores but unwind RNA helices through distinct mechanisms. Recent work has advanced our understanding of how their distinct mechanisms enable DEAD-box proteins to disrupt RNA base pairs on the surfaces of structured RNAs and RNPs, while some DEAH-box proteins are adept at disrupting base pairs in the interior of RNPs. Proteins from these families use these mechanisms to chaperone folding and promote rearrangements of structured RNAs and RNPs, including the spliceosome, and may use related mechanisms to maintain cellular mRNAs in unfolded or partially unfolded conformations.

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SARS‐CoV‐2 modulation of RIG‐I‐MAVS signaling: Potential mechanisms of impairment on host antiviral immunity and therapeutic approaches

Wang

Zhao

Liu

et al. 2022

MedComm – Future Medicine

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The coronavirus disease 2019 (COVID-19) is a global infectious disease aroused by RNA virus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Patients may suffer from severe respiratory failure or even die, posing a huge challenge to global public health. Retinoic acid-inducible gene I (RIG-I) is one of the major pattern recognition receptors, function to recognize RNA viruses and mediate the innate immune response. RIG-1 and melanoma differentiation-associated gene 5 contain an N-terminal caspase recruitment domain that is activated upon detection of viral RNA in the cytoplasm of virusinfected cells. Activated RIG-I and mitochondrial antiviral signaling (MAVS)protein trigger a series of corresponding immune responses such as the production of type I interferon against viral infection. In this review, we are summarizing the role of the structural, nonstructural, and accessory proteins from SARS-CoV-2 on the RIG-I-MAVS pathway, and exploring the potential mechanism how SARS-CoV-2 could evade the host antiviral response. We then proposed that modulation of the RIG-I-MAVS signaling pathway might be a novel and effective therapeutic strategy to against COVID-19 as well as the constantly mutating coronavirus.

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Structural insights into the mechanism of the DEAH-box RNA helicase Prp43

Cited by 90 publications

References 54 publications

The G-quadruplex (G4) resolvase DHX36 efficiently and specifically disrupts DNA G4s via a translocation-based helicase mechanism

The G-quadruplex (G4) resolvase DHX36 efficiently and specifically disrupts DNA G4s via a translocation-based helicase mechanism

Distinct RNA-unwinding mechanisms of DEAD-box and DEAH-box RNA helicase proteins in remodeling structured RNAs and RNPs

SARS‐CoV‐2 modulation of RIG‐I‐MAVS signaling: Potential mechanisms of impairment on host antiviral immunity and therapeutic approaches

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