Allosteric regulation of helicase core activities of the DEAD-box helicase YxiN by RNA binding to its RNA recognition motif

Samatanga, Brighton; Andreou, Alexandra Z.; Klostermeier, Dagmar

doi:10.1093/nar/gkx014

Cited by 12 publications

(15 citation statements)

References 50 publications

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“…Seemingly subtle differences in the helicase core sequence and structure between family members, such as altered degrees of interdomain communication, have drastic impacts on both thermodynamic interactions and the kinetics of duplex unwinding (Garbelli et al 2011;Samatanga and Klostermeier 2014). Amino and/or carboxy-terminal extensions can also influence enzymatic activity of individual members through various mechanisms, such as enhanced substrate selection, allosteric regulation, or nucleic acid tethering (Wang et al 2006;Banroques et al 2011;Mallam et al 2011;Rudolph and Klostermeier 2015;Samatanga et al 2017). Interestingly, 70% of DEAD-box protein genes in yeast are essential and provide nonredundant functions in vivo ranging from ribosome processing to translation and RNA decay (Rocak and Linder 2004).…”

Section: Introductionmentioning

confidence: 99%

Characterization of the mammalian DEAD-box protein DDX5 reveals functional conservation with S. cerevisiae ortholog Dbp2 in transcriptional control and glucose metabolism

Zheng

Wang

Tran

2017

RNA

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DEAD-box proteins are a class of nonprocessive RNA helicases that dynamically modulate the structure of RNA and ribonucleoprotein complexes (RNPs). However, the precise roles of individual members are not well understood. Work from our laboratory revealed that the DEAD-box protein Dbp2 in is an active RNA helicase in vitro that functions in transcription by promoting mRNP assembly, repressing cryptic transcription initiation, and regulating long noncoding RNA activity. Interestingly, Dbp2 is also linked to glucose sensing and hexose transporter gene expression. DDX5 is the mammalian ortholog of Dbp2 that has been implicated in cancer and metabolic syndrome, suggesting that the role of Dbp2 and DDX5 in glucose metabolic regulation is conserved. Herein, we present a refined biochemical and biological comparison of yeast Dbp2 and human DDX5 enzymes. We find that human DDX5 possesses a 10-fold higher unwinding activity than Dbp2, which is partially due to the presence of a mammalian/avian specific C-terminal extension. Interestingly, ectopic expression of rescues the cold sensitivity, cryptic initiation defects, and impaired glucose import in Δ cells, suggesting functional conservation. Consistently, we show that DDX5 promotes glucose uptake and glycolysis in mouse AML12 hepatocyte cells, suggesting that mammalian DDX5 and Dbp2 share conserved roles in cellular metabolism.

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Section: Introductionmentioning

confidence: 99%

Characterization of the mammalian DEAD-box protein DDX5 reveals functional conservation with S. cerevisiae ortholog Dbp2 in transcriptional control and glucose metabolism

Zheng

Wang

Tran

2017

RNA

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“…24,28 Modest allostery has also been demonstrated between the helicase core and a structured ancillary domain in the bacterial DEAD-box protein YxiN. 36 …”

Section: Discussionmentioning

confidence: 99%

The DEAD-Box Protein CYT-19 Uses Arginine Residues in Its C-Tail To Tether RNA Substrates

2017

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DEAD-box proteins are nonprocessive RNA helicases that play diverse roles in cellular processes. The Neurospora crassa DEAD-box protein CYT-19 promotes mitochondrial group I intron splicing and functions as a general RNA chaperone. CYT-19 includes a disordered, arginine-rich “C-tail” that binds RNA, positioning the helicase core to capture and unwind nearby RNA helices. Here we probed the C-tail further by varying the number and positions of arginines within it. We found that removing sets of as few as four of the 11 arginines reduced RNA unwinding activity (kcat/KM) to a degree equivalent to that seen upon removal of the C-tail, suggesting that a minimum or “threshold” number of arginines is required. In addition, a mutant with 16 arginines displayed RNA unwinding activity greater than that of wild-type CYT-19. The C-tail modifications impacted unwinding only of RNA helices within constructs that included an adjacent helix or structured RNA element that would allow C-tail binding, indicating that the helicase core remained active in the mutants. In addition, changes in RNA unwinding efficiency of the mutants were mirrored by changes in functional RNA affinity, as determined from the RNA concentration dependence of ATPase activity, suggesting that the C-tail functions primarily to increase RNA affinity. Interestingly, the salt concentration dependence of RNA unwinding activity is unaffected by C-tail composition, suggesting that the C-tail uses primarily hydrogen bonding, not electrostatic interactions, to bind double-stranded RNA. Our results provide insights into how an unstructured C-tail contributes to DEAD-box protein activity and suggest parallels with other families of RNA- and DNA-binding proteins.

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“…Hairpin 92 is part of the highly conserved peptidyl transferase center which remains unstructured until the final stages of 50S ribosomal subunit maturation (Nikolay et al 2018 ). The helicase activity of DbpA and YxiN is strongly activated upon binding of hairpin 92 to the RRM (Diges and Uhlenbeck 2001 ; Samatanga et al 2017 ), but the molecular mechanism behind this allosteric activation process is not clear.…”

Section: Biological Contextmentioning

confidence: 99%

Assignment of the Ile, Leu, Val, Met and Ala methyl group resonances of the DEAD-box RNA helicase DbpA from E. coli

Wurm

2020

Biomol NMR Assign

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ATP-dependent DEAD-box helicases constitute one of the largest families of RNA helicases and are important regulators of most RNA-dependent cellular processes. The functional core of these enzymes consists of two RecA-like domains. Changes in the interdomain orientation of these domains upon ATP and RNA binding result in the unwinding of double-stranded RNA. The DEAD-box helicase DbpA from E. coli is involved in ribosome maturation. It possesses a C-terminal RNA recognition motif (RRM) in addition to the canonical RecA-like domains. The RRM recruits DbpA to nascent ribosomes by binding to hairpin 92 of the 23S rRNA. To follow the conformational changes of Dbpa during the catalytic cycle we initiated solution state NMR studies. We use a divide and conquer approach to obtain an almost complete resonance assignment of the isoleucine, leucine, valine, methionine and alanine methyl group signals of full length DbpA (49 kDa). In addition, we also report the backbone resonance assignments of two fragments of DbpA that were used in the course of the methyl group assignment. These assignments are the first step towards a better understanding of the molecular mechanism behind the ATP-dependent RNA unwinding process catalyzed by DEAD-box helicases.

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Allosteric regulation of helicase core activities of the DEAD-box helicase YxiN by RNA binding to its RNA recognition motif

Cited by 12 publications

References 50 publications

Characterization of the mammalian DEAD-box protein DDX5 reveals functional conservation with S. cerevisiae ortholog Dbp2 in transcriptional control and glucose metabolism

Characterization of the mammalian DEAD-box protein DDX5 reveals functional conservation with S. cerevisiae ortholog Dbp2 in transcriptional control and glucose metabolism

The DEAD-Box Protein CYT-19 Uses Arginine Residues in Its C-Tail To Tether RNA Substrates

Assignment of the Ile, Leu, Val, Met and Ala methyl group resonances of the DEAD-box RNA helicase DbpA from E. coli

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