A single hydrophobic cleft in the Escherichia coli processivity clamp is sufficient to support cell viability and DNA damage-induced mutagenesis in vivo

Sutton, Mark D.; Duzen, Jill M.; Ponticelli, Sarah K. Scouten

doi:10.1186/1471-2199-11-102

Cited by 10 publications

(16 citation statements)

References 40 publications

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“…Importantly, a single cleft on the clamp was sufficient to support this switch, and both the rim and cleft contacts were required (19). Taken together, these findings suggest that Pol IV must first contact the rim of the ␤ clamp immediately adjacent to the cleft that is bound by Pol III* prior to gaining access to the same cleft previously associated with the stalled Pol III* (19,45) (Fig. 1).…”

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confidence: 54%

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Escherichia coli DNA Polymerase IV (Pol IV), but Not Pol II, Dynamically Switches with a Stalled Pol III* Replicase

et al. 2012

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The dnaN159 allele encodes a temperature-sensitive mutant form of the ␤ sliding clamp (␤159). SOS-induced levels of DNA polymerase IV (Pol IV) confer UV sensitivity upon the dnaN159 strain, while levels of Pol IV ϳ4-fold higher than those induced by the SOS response severely impede its growth. Here, we used mutations in Pol IV that disrupted specific interactions with the ␤ clamp to test our hypothesis that these phenotypes were the result of Pol IV gaining inappropriate access to the replication fork via a Pol III*-Pol IV switch relying on both the rim and cleft of the clamp. Our results clearly demonstrate that Pol IV relied on both the clamp rim and cleft interactions for these phenotypes. In contrast to the case for Pol IV, elevated levels of the other Pols, including Pol II, which was expressed at levels ϳ8-fold higher than the normal SOS-induced levels, failed to impede growth of the dnaN159 strain. These findings suggest that the mechanism used by Pol IV to switch with Pol III* is distinct from those used by the other Pols. Results of experiments utilizing purified components to reconstitute the Pol III*-Pol II switch in vitro indicated that Pol II switched equally well with both a stalled and an actively replicating Pol III* in a manner that was independent of the rim contact required by Pol IV. These results provide compelling support for the Pol III*-Pol IV two-step switch model and demonstrate important mechanistic differences in how Pol IV and Pol II switch with Pol III*.

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confidence: 54%

“…In addition to the cleft, Pols also contact noncleft surfaces (7,19,20,28,(42)(43)(44). These surfaces, together with a single cleft on the clamp, are sufficient for supporting viability of E. coli and managing the actions of its different Pols during UV-induced mutagenesis (45).…”

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confidence: 99%

Escherichia coli DNA Polymerase IV (Pol IV), but Not Pol II, Dynamically Switches with a Stalled Pol III* Replicase

et al. 2012

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“…As for Pol II, it was shown that the interaction of Pol IV with the b-clamp is essential to support its TLS activity in vivo (Becherel et al 2002). Until recently, it was thought that Pol III only contacts a single monomer of the dimeric b-clamp as shown in vitro (Sutton et al 2010). Together these observations appeared to comfort the so-called tool-belt model predicting simultaneous binding to the clamp of the replicative polymerase and a specialized polymerase (Pagès and Fuchs 2002;Indiani et al 2005).…”

Section: Rp Fuchs and S Fujiimentioning

confidence: 99%

Translesion DNA Synthesis and Mutagenesis in Prokaryotes

Fuchs

Fujii

2013

Cold Spring Harbor Perspectives in Biology

117

113

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The presence of unrepaired lesions in DNA represents a challenge for replication. Most, but not all, DNA lesions block the replicative DNA polymerases. The conceptually simplest procedure to bypass lesions during DNA replication is translesion synthesis (TLS), whereby the replicative polymerase is transiently replaced by a specialized DNA polymerase that synthesizes a short patch of DNA across the site of damage. This process is inherently error prone and is the main source of point mutations. The diversity of existing DNA lesions and the biochemical properties of Escherichia coli DNA polymerases will be presented. Our main goal is to deliver an integrated view of TLS pathways involving the multiple switches between replicative and specialized DNA polymerases and their interaction with key accessory factors. Finally, a brief glance at how other bacteria deal with TLS and mutagenesis is presented.

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“…We used nanoESI-MS under native conditions to assess whether sliding-clamp subunits from different species could form heterodimers with  Eco , with the rationale being that the dimerization interfaces would have to be chemically and structurally similar for formation of heterodimers, which would validate this interface as a broad-spectrum drug target. It is known that  Eco mutant subunits can be made to form heterodimers with the wild-type subunit (Scouten Ponticelli et al, 2009;Sutton et al, 2010;Jergic et al, 2013). Peaks in mass spectra corresponding to the m/z values of heterodimeric ions were observed for all pairwise combinations (Fig.…”

Section: Comparison Of Dimerization Interfacesmentioning

confidence: 89%

Crystal structures and biochemical characterization of DNA sliding clamps from three Gram-negative bacterial pathogens

McGrath

Martyn

Whittell

et al. 2018

Journal of Structural Biology

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Bacterial sliding clamps bind to DNA and act as protein-protein interaction hubs for several proteins involved in DNA replication and repair. The partner proteins all bind to a common pocket on sliding clamps via conserved linear peptide sequence motifs, which suggest the pocket as an attractive target for development of new antibiotics. Herein we report the X-ray crystal structures and biochemical characterization of β sliding clamps from the Gram-negative pathogens Pseudomonas aeruginosa, Acinetobacter baumannii and Enterobacter cloacae. The structures reveal close similarity between the pathogen and Escherichia coli clamps and similar patterns of binding to linear clamp-binding motif peptides. The results suggest that linear motifsliding clamp interactions are well conserved and an antibiotic targeting the sliding clamp should have broadspectrum activity against Gram-negative pathogens. Crystal structures and biochemical characterization of DNA sliding clamps from three Gramnegative bacterial pathogens, Journal of Structural Biology (2018), doi: https://doi.ABSTRACT Bacterial sliding clamps bind to DNA and act as protein-protein interaction hubs for several proteins involved in DNA replication and repair. The partner proteins all bind to a common pocket on sliding clamps via conserved linear peptide sequence motifs, which suggest the pocket as an attractive target for development of new antibiotics. Herein we report the X-ray crystal structures and biochemical characterization of  sliding clamps from the Gram-negative pathogens Pseudomonas aeruginosa, Acinetobacter baumannii and Enterobacter cloacae. The structures reveal close similarity between the pathogen and Escherichia coli clamps and similar patterns of binding to linear clamp-binding motif peptides. The results suggest that linear motif-sliding clamp interactions are well conserved and an antibiotic targeting the sliding clamp should have broadspectrum activity against Gram-negative pathogens.

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A single hydrophobic cleft in the Escherichia coli processivity clamp is sufficient to support cell viability and DNA damage-induced mutagenesis in vivo

Cited by 10 publications

References 40 publications

Escherichia coli DNA Polymerase IV (Pol IV), but Not Pol II, Dynamically Switches with a Stalled Pol III* Replicase

Escherichia coli DNA Polymerase IV (Pol IV), but Not Pol II, Dynamically Switches with a Stalled Pol III* Replicase

Translesion DNA Synthesis and Mutagenesis in Prokaryotes

Crystal structures and biochemical characterization of DNA sliding clamps from three Gram-negative bacterial pathogens

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