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Literature summary for 3.6.4.B7 extracted from

  • Torres, R.; Serrano, E.; Alonso, J.
    Bacillus subtilis RecA interacts with and loads RadA/Sms to unwind recombination intermediates during natural chromosomal transformation (2019), Nucleic Acids Res., 47, 9198-9215 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 3.6.4.B7

Activating Compound

Activating Compound Comment Organism Structure
RecA RecA interacts with and loads wild-type RadA to promote unwinding of a non-cognate 3'-tailed or 5'-fork DNA substrate and of a 5'-invading D-loop. Wild-type RadA interaction with RecA is crucial to recruit the former onto D-loop DNA, and both proteins in concert catalyse D-loop extension to favour integration of ssDNA during chromosomal transformation. Proposed model for the action of the 5'->3' RadA helicase in coordination with RecA during natural transformation and in doublee strand break repair, overview. RecA has ATPase activity, but RadA inhibits the ATPase activity of RecA. Mutational function analysis, overview Bacillus subtilis

Cloned(Commentary)

Cloned (Comment) Organism
gene radA, recombinant enzyme expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)[pLysS] from plasmid pCB1020 under the control of a rifampicin-resistant promoter, PT7 Bacillus subtilis

Protein Variants

Protein Variants Comment Organism
C13A site-directed mutagenesis of the tetracysteine motif, the mutant variant binds ssDNA, and this interaction stimulates its ATPase activity. Wild-type RadA interacts with and inhibits the ATPase activity of RecA, but mutant RadA C13A fails to do so Bacillus subtilis
C13R site-directed mutagenesis of the tetracysteine motif, the mutant variant binds ssDNA, and this interaction stimulates its ATPase activity Bacillus subtilis
K104R site-directed mutagenesis, the mutant variant in the Walker A (radA1041 [K104R]) motif forms a complex with RecA Bacillus subtilis

Localization

Localization Comment Organism GeneOntology No. Textmining

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Bacillus subtilis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + H2O Bacillus subtilis
-
 ADP + phosphate
-
 ?
ATP + H2O Bacillus subtilis 168
-
 ADP + phosphate
-
 ?

Organism

Organism UniProt Comment Textmining
Bacillus subtilis P37572
-
-
Bacillus subtilis 168 P37572
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3)[pLysS] by nickel affinity chromatography Bacillus subtilis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + H2O
-
 Bacillus subtilis ADP + phosphate
-
 ?
ATP + H2O
-
 Bacillus subtilis 168 ADP + phosphate
-
 ?
additional information performance of transformation and survival assays, DNA helicase assays, ATP hydrolysis assays, and protein-DNA or protein-protein interactions analysis Bacillus subtilis ?
-
-
additional information performance of transformation and survival assays, DNA helicase assays, ATP hydrolysis assays, and protein-DNA or protein-protein interactions analysis Bacillus subtilis 168 ?
-
-

Subunits

Subunits Comment Organism
? x * 49400, recombinant enzyme, SDS-PAGE Bacillus subtilis
More RadA has four well-conserved motifs: a potential C4-type zinc-binding motif at the N-terminal domain, a central canonical RecA-like ATPase domain (H1-H4 motifs) and KNRFG motif, and the P/LonC domain at the C-terminus domain Bacillus subtilis

Synonyms

Synonyms Comment Organism
RadA
-
 Bacillus subtilis
SMS
-
 Bacillus subtilis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Bacillus subtilis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
 assay at Bacillus subtilis

General Information

General Information Comment Organism
evolution Bacillus subtilis encodes three branch migration translocases: RuvAB, RecG, and RadA, i.e. Sms Bacillus subtilis
malfunction a null radA mutation impairs chromosomal transformation, in the absence of RadA competent cells require the RecG translocase for natural chromosomal transformation. A RadA/SmsC4 mutation impairs chromosomal and plasmid transformation. Enzyme mutants RadA C13A or C13R fail to interact with RecA and do not promote unwinding of a non-cognate 3'-tailed or 5'-fork DNA substrate. Enzyme mutants RadA C13A and C13R hydrolyse ATP in a ssDNA-dependent manner. Mutant RadA C13A interacts with itself but does not interact with RecA. RadA C13A and C13R variants preferentially bind ssDNA, albeit with lower efficiency than the wild-type enzyme. RadA C13A and C13R mutants bind natural ssDNA and partially displace SsbA Bacillus subtilis
physiological function wild-type RadA interacts with and inhibits the ATPase activity of RecA (BG214). RadA and its mutant variants, C13A and C13R, bound to the 5'-tail of a DNA substrate, unwind DNA in the 5'->3' direction. RecA interacts with and loads wild-type RadA to promote unwinding of a non-cognate 3'-tailed or 5'-fork DNA substrate. Wild-type RadA interaction with RecA is crucial to recruit the former onto D-loop DNA, and both proteins in concert catalyse D-loop extension to favour integration of ssDNA during chromosomal transformation. But RadA inhibits the ATPase activity of RecA. Proposed model for the action of the 5'->3' RadA helicase in coordination with RecA during natural transformation and in double strand break repair, overview. RadA is crucial for chromosomal transformation, but is essential in the DELTArecG background. Functional analysis, detailed overview Bacillus subtilis