Recombinant Pyrococcus furiosus DNA repair and recombination protein radB (radB)
Description
Functional Roles in DNA Repair and Recombination
RadB participates in homologous recombination (HR) but exhibits distinct biochemical activities compared to its paralog RadA:
RadB’s primary role involves modulating recombination intermediates rather than catalyzing strand exchange directly .
Interaction Networks
RadB forms critical partnerships with multiple DNA repair proteins:
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PolD (DNA polymerase II): Binds the DP1 subunit via residues 206–498 of DP1, overlapping a conserved region essential for PolD activity .
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RadA: Co-precipitates with RadA, suggesting collaborative roles in HR .
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Hjc resolvase: Inhibits Holliday junction cleavage by Hjc in an ATP-dependent manner .
Table 1: RadB interaction partners and functional outcomes
Biochemical Characterization
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ATPase activity: RadB hydrolyzes ATP at 0.2% the rate of RadA, with no DNA stimulation .
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DNA binding: Exhibits high affinity for both single-stranded (ssDNA) and double-stranded DNA (dsDNA) .
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Recombinant production: Expressed in Escherichia coli and insect cells, purified via affinity chromatography (e.g., GST-tagged constructs) .
Genetic and Cellular Roles
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Recombination efficiency: ΔradB strains show reduced HR efficiency and increased sensitivity to DNA damage .
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Regulatory function: Acts as a recombination mediator, promoting RadA filament formation on DNA . Suppressor mutations in radA (e.g., L126P, G384D) restore viability in ΔradB strains, implicating RadB in RadA activation .
Regulatory Mechanisms
RadB’s activity is modulated by ATP:
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ATP absence: Stabilizes RadB-Hjc complexes, inhibiting Hjc cleavage .
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ATP presence: Dissociates RadB from Hjc, restoring resolvase activity .
Research Implications
RadB’s dual roles in bridging replication/recombination and regulating Holliday junction resolution highlight its importance in maintaining genome stability under extreme conditions. Its mediator function parallels eukaryotic Rad51 paralogs (e.g., Rad55-Rad57), suggesting conserved regulatory principles across domains .
Product Specs
Target Background
KEGG: pfu:PF0021
STRING: 186497.PF0021
Q&A
Basic Research Questions
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What is RadB protein from Pyrococcus furiosus?
RadB is a DNA repair and recombination protein found in the hyperthermophilic archaeon Pyrococcus furiosus. It is a paralogue of RadA, which is the archaeal homolog of the eukaryotic Rad51 and bacterial RecA recombinases . The full-length RadB protein consists of 220 amino acids and plays a role in homologous recombination processes, particularly as a recombination mediator assisting in RadA-mediated strand exchange . RadB belongs to the RecA-family of recombinases, which are central to the repair of double-strand DNA breaks, the restart of stalled replication forks, and the generation of genetic diversity .
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What is the proposed function of RadB in archaea?
The primary function of RadB in archaea is proposed to be a recombination mediator, assisting in RadA-mediated strand exchange during homologous recombination . Studies in Haloferax volcanii have shown that RadB is required for efficient recombination and survival following treatment with DNA-damaging agents . The mechanism appears to involve inducing a conformational change in RadA and thereby promoting its polymerization on DNA, which is essential for the strand exchange process that underlies homologous recombination . This mediator role positions RadB as a key regulatory component in archaeal DNA repair systems, though its importance seems to vary across different archaeal species.
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How does the requirement for RadB differ between archaeal species?
The requirement for RadB varies significantly between archaeal species. In Haloferax volcanii, RadB is required for efficient recombination and survival following DNA damage, with deletion strains showing marked phenotypes . In contrast, deletion of radB in Pyrococcus furiosus strain COM1 had no apparent effect on either recombination or DNA transformation . This suggests that the role of RadB may be species-specific or that functional redundancy exists in some archaeal systems but not others. These differences may reflect adaptations to different environmental niches or fundamental differences in recombination mechanisms between archaeal lineages. The varying requirement highlights the diversity of recombination mechanisms even within the archaeal domain.
