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Information on EC 3.6.4.13 - RNA helicase and Organism(s) Escherichia coli and UniProt Accession P0A9P6

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EC Tree
IUBMB Comments
RNA helicases utilize the energy from ATP hydrolysis to unwind RNA. Some of them unwind RNA with a 3' to 5' polarity , other show 5' to 3' polarity . Some helicases unwind DNA as well as RNA [7,8]. May be identical with EC 3.6.4.12 (DNA helicase).
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This record set is specific for:
Escherichia coli
UNIPROT: P0A9P6
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The taxonomic range for the selected organisms is: Escherichia coli
The enzyme appears in selected viruses and cellular organisms
Synonyms
helicase, rig-i, rna helicase, eif4a, ddx3x, dead-box rna helicase, ns3 helicase, dead-box helicase, ddx21, rna helicase a, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ATP-dependent RNA helicase
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cold-shock DEAD-box protein A
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DEAD-box RNA helicase
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RNA helicase CsdA
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DEAD-box RNA helicase
DEAD-box rRNA helicase
-
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helicase B
-
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
ATP + H2O = ADP + phosphate
show the reaction diagram
SYSTEMATIC NAME
IUBMB Comments
ATP phosphohydrolase (RNA helix unwinding)
RNA helicases utilize the energy from ATP hydrolysis to unwind RNA. Some of them unwind RNA with a 3' to 5' polarity [3], other show 5' to 3' polarity [8]. Some helicases unwind DNA as well as RNA [7,8]. May be identical with EC 3.6.4.12 (DNA helicase).
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + H2O
ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + H2O
ADP + phosphate
show the reaction diagram
RNA + H2O
?
show the reaction diagram
-
helicase/unwinding activity
-
-
?
additional information
?
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + H2O
ADP + phosphate
show the reaction diagram
-
-
-
?
ATP + H2O
ADP + phosphate
show the reaction diagram
RNA + H2O
?
show the reaction diagram
-
helicase/unwinding activity
-
-
?
additional information
?
-
-
helicase B, RhlB, is one of the five DEAD box RNA-dependent ATPases in Escherichia coli. ATPases found in Escherichia coli. RhlB requires an interaction with the partner protein RNase E for appreciable ATPase and RNA unwinding activities
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-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
RNase E
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is required for ATPase and RNA unwinding activities of the enzyme, forms a complex with the enzyme, interaction analysis, overview. Avid, enthalpy-favored interaction between the helicase and RNase E 696-762 with an equilibrium binding constant Kaof at least 1 x 108 M-1 determined by isothermal titration calorimetry. Protein-protein and RNA-binding surfaces both communicate allosterically with the ATPase catalytic center
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rRNA
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activates the ATPase activity of DbpA by promoting a conformational change after ATP binding that is associated with hydrolysis
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
Michaelis-Menten kinetics of ATP hydrolysis
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.72 - 1.28
ATP
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.00000001
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about, wild-type enzyme, RNA helicase activity
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
UniProt
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
DEAD-box proteins belong to a ubiquitous family of RNA helicases, which are widely found from prokaryotes to eukaryotes and participate in multiple cellular processes, such as premRNA splicing, translation initiation, modulating RNA-protein complexes, RNA decay, and ribosome biogenesis. Sequence alignment and architecture of different DEAD-Box proteins, overview
malfunction
breaking the sequence of the interdomain peptide linker and inserting the 23 amino acids peptide segment causes a decrease in binding affinity, likely as a consequence of formation of non-native interaction between the insert peptide and the RNA molecule or other regions of the protein and not a consequence of disrupting native interactions between the DbpA RNA binding domain and the interdomain linker. The peptide extension is not effecting the formation of the proper ATP pocket, but the ATP turnover rate is affected by the peptide extension. Although the ATP turnover of the extended DbpA is reduced when compared to wild-type DbpA, extended DbpA is a much more efficient enzyme than many members of DEAD-box family of proteins. The reduction on the ATP turnover of the extended DbpA is a consequence of its decrease in binding affinity for RNA. The extension of the interdomain linker region has no effect on the ability of DbpA to perform its helicase function. Thus, the physical connection of DbpA RNA binding domain to the catalytic core is unimportant for the helicase activity of DbpA, suggesting the DbpA protein is a region-specific enzyme, which would unwind any double-helix substrate near hairpin 92
physiological function
DbpA is a DEAD-box RNA helicase implicated in RNA structural rearrangements in the peptidyl transferase center. DbpA performs RNA structural isomerizations in a region of the ribosome that is evolutionarily conserved in all organisms and crucial for their survival
additional information
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
the helicase core of CsdA is comprised of two RecA-like domains (RecA1 and RecA2) joined by a flexible linker and contains all conserved motifs, two previously auxiliary domains are found: a dimerization domain (DD) and an RNA-binding domain (RBD), conformational flexibilities of the helicase core domains and C-terminal regions, enzyme domain structure, three-dimensional modelling, detailed overview. DD is indispensable for stabilizing the CsdA dimeric structure. Structure comparisons
monomer
-
DbpA is monomeric in solution up to a concentration of 25 mM and over the temperature range of 4°C to 22°C
additional information
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location of the nine conserved sequence motifs in the DEAD box helicase RhlB, structure modelling, overview
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
structure of CsdA_218-445 is determined by X-ray diffraction and refined to an R factor of 0.225, with an Rfree of 0.268 at a resolution of 2.3 A by molecular replacement using Hera (PDB ID 3EAQ) as the model. The structure of CsdA_218-445 includes two RecA-like domains (RecA2) and two DDs, which form a V-shape dimer. The V-shape conformation of the CsdA_218-445 dimer in solution is further confirmed by SAXS experiments. Conformational flexibilities of CsdA_1-445 and CsdA_FL are revealed by SAXS
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D310H
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site-directed mutagenesis of the V motif, leads to altered enzyme activity, overview
D313H
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site-directed mutagenesis of the V motif, leads to altered enzyme activity, overview
H320D
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site-directed mutagenesis of the V motif, leads to altered enzyme activity, overview
Y383A
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site-directed mutagenesis, the mutation causes the formation of a higher order molecular weight species in binding of RNaseE by RhlB
additional information
in order to increase the peptide linker region length, a 23 amino acid residue polypeptide with a composition of NASSGSSASSPSASNSPGANGSS was inserted between the native interdomain region's Ala and Thr residues. The sequence of the new extended interdomain linker is PANSSIANASSGSSASSPSASNSPGANGSSTLEAE. This peptide sequence is chosen because it has similar structural and dynamic properties to the native interdomain region, and both the designed and the native interdomain linker are predicted to form a flexible and unstructured region. In addition, since DbpA is purified as a native protein, small and polar amino acids, which promote peptide solubility, are placed into the polypeptide insert to discourage the aggregation of extended DbpA and its partition into inclusion bodies. The new interdomain linker is not digested by the Escherichia coli proteolytic enzymes and the extended DbpA is expressed as an intact and soluble protein. Breaking the sequence of the interdomain peptide linker and inserting the 23 amino acids peptide segment causes a decrease in binding affinity, likely as a consequence of formation of non-native interaction between the insert peptide and the RNA molecule or other regions of the protein and not a consequence of disrupting native interactions between the DbpA RNA binding domain and the interdomain linker. The peptide extension is not effecting the formation of the proper ATP pocket, but the ATP turnover rate is affected by the peptide extension. Although the ATP turnover of the extended DbpA is reduced when compared to wild-type DbpA, extended DbpA is a much more efficient enzyme than many members of DEAD-box family of proteins. The reduction on the ATP turnover of the extended DbpA is a consequence of its decrease in binding affinity for RNA. The extension of the interdomain linker region has no effect on the ability of DbpA to perform its helicase function. Thus, the physical connection of DbpA RNA binding domain to the catalytic core is unimportant for the helicase activity of DbpA, suggesting the DbpA protein is a region-specific enzyme, which would unwind any double-helix substrate near hairpin 92
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged wild-type and mutant RhlB and RNaseE from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration
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recombinant N-terminally His-tagged wild-type and engineered enzymes from Escherichia coli by nickel affinity chromatography and gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
construction of a di-cistronic vector that overexpresses a complex comprising RhlB and its recognition site within RNase E, corresponding to residues 696-762, the expression construct is termed pRneRhlBDELTA1-397. Expression of His-tagged wild-type and mutant RhlB and RNaseE in Escherichia coli strain BL21(DE3)
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recombinant expression of N-terminally His-tagged wild-type and engineered enzymes in Escherichia coli
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Diges, C.M.; Uhlenbeck, O.C.
Escherichia coli DbpA is a 3' -> 5' RNA helicase
Biochemistry
31; 44
7903-7911
2005
Escherichia coli
Manually annotated by BRENDA team
Talavera, M.A.; Matthews, E.E.; Eliason, W.K.; Sagi, I.; Wang, J.; Henn, A.; De La Cruz, E.M.
Hydrodynamic characterization of the DEAD-box RNA helicase DbpA
J. Mol. Biol.
355
697-707
2005
Escherichia coli
Manually annotated by BRENDA team
Henn, A.; Cao, W.; Hackney, D.D.; De La Cruz, E.M.
The ATPase cycle mechanism of the DEAD-box rRNA helicase, DbpA
J. Mol. Biol.
377
193-205
2008
Escherichia coli
Manually annotated by BRENDA team
Worrall, J.A.; Howe, F.S.; McKay, A.R.; Robinson, C.V.; Luisi, B.F.
Allosteric activation of the ATPase activity of the Escherichia coli RhlB RNA helicase
J. Biol. Chem.
283
5567-5576
2008
Escherichia coli
Manually annotated by BRENDA team
Moore, A.F.; Gentry, R.C.; Koculi, E.
DbpA is a region-specific RNA helicase
Biopolymers
107
e23001
2017
Escherichia coli (P21693)
Manually annotated by BRENDA team
Xu, L.; Wang, L.; Peng, J.; Li, F.; Wu, L.; Zhang, B.; Lv, M.; Zhang, J.; Gong, Q.; Zhang, R.; Zuo, X.; Zhang, Z.; Wu, J.; Tang, Y.; Shi, Y.
Insights into the structure of dimeric RNA helicase CsdA and indispensable role of its C-terminal regions
Structure
25
1795-1808.e5
2017
Escherichia coli (P0A9P6)
Manually annotated by BRENDA team