Structural basis for DNA 5´-end resection by RecJ

Cheng, Kaiying; Xu, Hong; Chen, Xuanyi; Wang, Liangyan; Teng, Bing; Zhao, Ye; Hua, Yuejin

doi:10.7554/elife.14294

Cited by 59 publications

(102 citation statements)

References 62 publications

(92 reference statements)

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“…RecJ, GAN, and Cdc45 are characterized by two catalytic domains, the N-terminal DHH domain and the following DHH1 domain, with seven conserved motifs, I to IV on DHH and VI and VII (or DHHA1 motif) on DHH1, and the active site is formed between the two domains (28–30). Cdc45 lacks nuclease activity, due to replacements of the catalytic residues (Supplementary Figure S1).…”

Section: Resultsmentioning

confidence: 99%

“…The active site of RecJ/GAN is formed by several key residues on the conserved motifs of DHH, which coordinates one or two divalent metal ions such as Mg 2+ or Mn 2+ to stabilize the scissile phosphate and activate the nucleophilic-attacking water (28–30). These amino acid residues are conserved in GAN, suggesting that it cleaves DNA in a similar manner to other RecJ enzymes (Supplementary Figure S1).…”

Section: Resultsmentioning

confidence: 99%

“…The crystal structure of DraRecJ has been solved in complexes with a mononucleotide or an ssDNA, which provided the detailed interactions between the enzyme and DNA (30). A structural comparison between this bacterial enzyme–DNA complex with GAN indicated that the DNA binding mode of GAN is partly similar to that of DraRecJ, particularly around DHH and DHH1.…”

Section: Resultsmentioning

confidence: 99%

“…First, an archaeal GINS from Thermococcus kodakaraensis stimulated the activity of its cognate exonuclease (TK_RS06185), which was designated as GAN (GINS-associated nuclease) (26). Second, the crystal structure of human Cdc45 (27) partly resembles those of the bacterial RecJ enzymes from Thermus thermophilus (28,29) and Deinococcus radiodurans (30). …”

Section: Introductionmentioning

confidence: 99%

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Atomic structure of an archaeal GAN suggests its dual roles as an exonuclease in DNA repair and a CMG component in DNA replication

Oyama

Ishino

Shirai

et al. 2016

Nucleic Acids Research

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In eukaryotic DNA replication initiation, hexameric MCM (mini-chromosome maintenance) unwinds the template double-stranded DNA to form the replication fork. MCM is activated by two proteins, Cdc45 and GINS, which constitute the ‘CMG’ unwindosome complex together with the MCM core. The archaeal DNA replication system is quite similar to that of eukaryotes, but only limited knowledge about the DNA unwinding mechanism is available, from a structural point of view. Here, we describe the crystal structure of an archaeal GAN (GINS-associated nuclease) from Thermococcus kodakaraensis, the homolog of eukaryotic Cdc45, in both the free form and the complex with the C-terminal domain of the cognate Gins51 subunit (Gins51C). This first archaeal GAN structure exhibits a unique, ‘hybrid’ structure between the bacterial RecJ and the eukaryotic Cdc45. GAN possesses the conserved DHH and DHH1 domains responsible for the exonuclease activity, and an inserted CID (CMG interacting domain)-like domain structurally comparable to that in Cdc45, suggesting its dual roles as an exonuclease in DNA repair and a CMG component in DNA replication. A structural comparison of the GAN–Gins51C complex with the GINS tetramer suggests that GINS uses the mobile Gins51C as a hook to bind GAN for CMG formation.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Atomic structure of an archaeal GAN suggests its dual roles as an exonuclease in DNA repair and a CMG component in DNA replication

Oyama

Ishino

Shirai

et al. 2016

Nucleic Acids Research

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“…RecJ, which functions in a RecFOR pathway of DNA break repair, belongs to the DHH phosphodiesterase enzyme superfamily (7). M. smegmatis HR in the absence of the AdnAB nucleases requires RecO and RecR (4).…”

mentioning

confidence: 99%

The DNA Repair Repertoire of Mycobacterium smegmatis FenA Includes the Incision of DNA 5′ Flaps and the Removal of 5′ Adenylylated Products of Aborted Nick Ligation

2017

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We characterize Mycobacterium smegmatis FenA as a manganese-dependent 5=-flap endonuclease homologous to the 5=-exonuclease of DNA polymerase I. FenA incises a nicked 5= flap between the first and second nucleotides of the duplex segment to yield a 1-nucleotide gapped DNA, which is then further resected in dinucleotide steps. Initial FenA cleavage at a Y-flap or nick occurs between the first and second nucleotides of the duplex. However, when the template 3= single strand is eliminated to create a 5=-tailed duplex, FenA incision shifts to between the second and third nucleotides. A double-flap substrate with a mobile junction (mimicking limited strand displacement synthesis during gap repair) is preferentially incised as the 1-nucleotide 3=-flap isomer, with the scissile phosphodiester shifted by one nucleotide versus a static double flap. FenA efficiently removes the 5= App(dN) terminus of an aborted nick ligation reaction intermediate, thereby highlighting FenA as an agent of repair of such lesions, which are formed under a variety of circumstances by bacterial NAD ϩ -dependent DNA ligases and especially by mycobacterial DNA ligases D and C.IMPORTANCE Structure-specific DNA endonucleases are implicated in bacterial DNA replication, repair, and recombination, yet there is scant knowledge of the roster and catalytic repertoire of such nucleases in Mycobacteria. This study identifies M. smegmatis FenA as a stand-alone endonuclease homologous to the 5=-exonuclease domain of mycobacterial DNA polymerase 1. FenA incises 5= flaps, 5= nicks, and 5= App(dN) intermediates of aborted nick ligation. The isolated N-terminal domain of M. smegmatis Pol1 is also shown to be a flap endonuclease.KEYWORDS DNA repair, flap endonuclease, mycobacteria H omologous recombination (HR) in mycobacteria initiates via resection of a DNA double-strand break (DSB) by the AdnAB helicase-nuclease to generate a 3= single-strand DNA substrate for RecA-mediated strand invasion. AdnAB consists of two subunits, AdnA and AdnB, each composed of an N-terminal ATPase domain and a C-terminal nuclease domain (1-3). DSB unwinding by AdnAB in vitro is stringently dependent on the ATPase activity of the AdnB motor translocating on the 3= DNA strand. Unwinding drives the 3= DNA strand into the AdnB nuclease domain and threads the 5= DNA strand into the AdnA nuclease domain. Recent genetic analyses in Mycobacterium smegmatis revealed that a nuclease-dead AdnAB enzyme can sustain mycobacterial HR in vivo, as long as its AdnB motor is intact (4). Thus, AdnAB's processive DSB unwinding activity suffices for AdnAB function in HR. These findings raised two possibilities: (i) mycobacteria have a backup nuclease that resects the 5= DNA strand unwound by AdnAB, or (ii) HR can proceed via DSB unwinding and capture of

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The mechanism of Single strand binding protein–RecG binding: Implications for SSB interactome function

et al. 2020

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The Escherichia coli single-strand DNA binding protein (SSB) is essential to viability where it functions to regulate SSB interactome function. Here it binds to single-stranded DNA and to target proteins that comprise the interactome. The region of SSB that links these two essential protein functions is the intrinsically disordered linker. Key to linker function is the presence of three, conserved PXXP motifs that mediate binding to oligosaccharide-oligonucleotide binding folds (OB-fold) present in SSB and its interactome partners. Not surprisingly, partner OB-fold deletions eliminate SSB binding. Furthermore, single point mutations in either the PXXP motifs or, in the RecG OB-fold, obliterate SSB binding. The data also demonstrate that, and in contrast to the view currently held in the field, the C-terminal acidic tip of SSB is not required for interactome partner binding. Instead, we propose the tip has two roles.First, and consistent with the proposal of Dixon, to regulate the structure of the C-terminal domain in a biologically active conformation that prevents linkers from binding to SSB OB-folds until this interaction is required. Second, as a secondary binding domain. Finally, as OB-folds are present in SSB and many of its partners, we present the SSB interactome as the first family of OB-fold genome guardians identified in prokaryotes. K E Y W O R D SDNA helicase, OB-fold, PXXP motif, RecG, SH3 domain, single-strand binding protein, SSB interactome

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Structural basis for DNA 5´-end resection by RecJ

Cited by 59 publications

References 62 publications

Atomic structure of an archaeal GAN suggests its dual roles as an exonuclease in DNA repair and a CMG component in DNA replication

Atomic structure of an archaeal GAN suggests its dual roles as an exonuclease in DNA repair and a CMG component in DNA replication

The DNA Repair Repertoire of Mycobacterium smegmatis FenA Includes the Incision of DNA 5′ Flaps and the Removal of 5′ Adenylylated Products of Aborted Nick Ligation

The mechanism of Single strand binding protein–RecG binding: Implications for SSB interactome function

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