Dynamics of Open DNA Sliding Clamps

Oakley, Aaron J.

doi:10.1371/journal.pone.0154899

Cited by 24 publications

(24 citation statements)

References 53 publications

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“…Sliding clamps are present in all organisms: bacteria, eukaryotes and viruses ( 10 ). In this study, we looked for similarities and differences in the dynamics of four different but similarly structured ring-shaped proteins on DNA: sliding clamps from three different organisms (the eukaryotic PCNA, the prokaryotic β-clamp, and the virus gp45 protein) and the 9-1-1 repair protein (see Figures 6 and 7 ).…”

Section: Resultsmentioning

confidence: 99%

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Diffusion of ring-shaped proteins along DNA: case study of sliding clamps

Daitchman

Greenblatt

Levy

2018

Nucleic Acids Research

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Several DNA-binding proteins, such as topoisomerases, helicases and sliding clamps, have a toroidal (i.e. ring) shape that topologically traps DNA, with this quality being essential to their function. Many DNA-binding proteins that function, for example, as transcription factors or enzymes were shown to be able to diffuse linearly (i.e. slide) along DNA during the search for their target binding sites. The protein's sliding properties and ability to search DNA, which often also involves hopping and dissociation, are expected to be different when it encircles the DNA. In this study, we explored the linear diffusion of four ring-shaped proteins of very similar structure: three sliding clamps (PCNA, β-clamp, and the gp45) and the 9-1-1 protein, with a particular focus on PCNA. Coarse-grained molecular dynamics simulations were performed to decipher the sliding mechanism adopted by these ring-shaped proteins and to determine how the molecular properties of the inner and outer ring govern its search speed. We designed in silico variants to dissect the contributions of ring geometry and electrostatics to the sliding speed of ring-shaped proteins along DNA. We found that the toroidal proteins diffuse when they are tilted relative to the DNA axis and able to rotate during translocation, but that coupling between rotation and translocation is quite weak. Their diffusion speed is affected by the shape of the inner ring and, to a lesser extent, by its electrostatic properties. However, breaking the symmetry of the electrostatic potential can result in deviation of the DNA from the center of the ring and cause slower linear diffusion. The findings are discussed in light of earlier computational and experimental studies on the sliding of clamps.

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Section: Resultsmentioning

confidence: 99%

“…The sliding clamps of eukaryotes and archaea are called PCNA (Proliferating Cell Nuclear Antigen) ( 7 ), those of bacteria are the β-clamps (i.e. the β subunit of pol III) ( 8 , 9 ), and the clamp of bacteriophage T4 is called gp45 ( 10 ). All clamps are homomers: the PCNA and gp45 are trimers while the β-clamp is a dimer.…”

Section: Introductionmentioning

confidence: 99%

Diffusion of ring-shaped proteins along DNA: case study of sliding clamps

Daitchman

Greenblatt

Levy

2018

Nucleic Acids Research

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“…Similarly, a low resolution (12 Å) cryo-electron microscopy structure of archaeal PCNA in complex with RFC and DNA from Pyrococcus furiosus also reveals out-of-plane ring-opening ( 78 ). However, biochemical and computational evidence suggest that sliding DNA clamps may sample ring-open and ring-closed conformations ( 29 , 79 ), even in the absence of their respective clamp loaders, despite the aforementioned structural data. In fact, the ring-open conformation of gp45 was shown to dominate in solution by fluorescence spectroscopy ( 29 ), while only the ring-closed structure (in the presence of clamp loader) could be solved by X-ray crystallography ( 30 ).…”

Section: Discussionmentioning

confidence: 99%

Investigation of sliding DNA clamp dynamics by single-molecule fluorescence, mass spectrometry and structure-based modeling

Gadkari

Harvey

Raper

et al. 2018

Nucleic Acids Research

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Proliferating cell nuclear antigen (PCNA) is a trimeric ring-shaped clamp protein that encircles DNA and interacts with many proteins involved in DNA replication and repair. Despite extensive structural work to characterize the monomeric, dimeric, and trimeric forms of PCNA alone and in complex with interacting proteins, no structure of PCNA in a ring-open conformation has been published. Here, we use a multidisciplinary approach, including single-molecule Förster resonance energy transfer (smFRET), native ion mobility-mass spectrometry (IM-MS), and structure-based computational modeling, to explore the conformational dynamics of a model PCNA from Sulfolobus solfataricus (Sso), an archaeon. We found that Sso PCNA samples ring-open and ring-closed conformations even in the absence of its clamp loader complex, replication factor C, and transition to the ring-open conformation is modulated by the ionic strength of the solution. The IM-MS results corroborate the smFRET findings suggesting that PCNA dynamics are maintained in the gas phase and further establishing IM-MS as a reliable strategy to investigate macromolecular motions. Our molecular dynamic simulations agree with the experimental data and reveal that ring-open PCNA often adopts an out-of-plane left-hand geometry. Collectively, these results implore future studies to define the roles of PCNA dynamics in DNA loading and other PCNA-mediated interactions.

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“…As a result, Pol V plays a greater role in bypassing MMS-induced lesions, explaining the ~2- to ~3-fold increase in mutation frequency ( Fig 5A ). Although G22 of the β clamp failed to contact DNA in the crystal [ 27 ], flexibility at this position may contribute to the ability of R24 to interact with DNA [ 60 – 62 ]. In this case, the modest Pol IV defect observed for dnaN-G22A and dnaN-R24A could result from impaired β clamp-DNA interactions.…”

Section: Discussionmentioning

confidence: 99%

Identification of β Clamp-DNA Interaction Regions That Impair the Ability of E. coli to Tolerate Specific Classes of DNA Damage

et al. 2016

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The E. coli dnaN-encoded β sliding clamp protein plays a pivotal role in managing the actions on DNA of the 5 bacterial DNA polymerases, proteins involved in mismatch repair, as well as several additional proteins involved in DNA replication. Results of in vitro experiments indicate that the loading of β clamp onto DNA relies on both the DnaX clamp loader complex as well as several discrete sliding clamp-DNA interactions. However, the importance of these DNA interactions to E. coli viability, as well as the ability of the β clamp to support the actions of its numerous partner proteins, have not yet been examined. To determine the contribution of β clamp-DNA interactions to the ability of E. coli to cope with different classes of DNA damage, we used alanine scanning to mutate 22 separate residues mapping to 3 distinct β clamp surfaces known or nearby those known to contact the DNA template, including residues P20-L27 (referred to here as loop I), H148-Y154 (loop II) and 7 different residues lining the central pore of the β clamp through which the DNA template threads. Twenty of these 22 dnaN mutants supported bacterial growth. While none of these 20 conferred sensitivity to hydrogen peroxide or ultra violet light, 12 were sensitized to NFZ, 5 were sensitized to MMS, 8 displayed modestly altered frequencies of DNA damage-induced mutagenesis, and 2 may be impaired for supporting hda function. Taken together, these results demonstrate that discrete β clamp-DNA interaction regions contribute to the ability of E. coli to tolerate specific classes of DNA damage.

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Dynamics of Open DNA Sliding Clamps

Cited by 24 publications

References 53 publications

Diffusion of ring-shaped proteins along DNA: case study of sliding clamps

Diffusion of ring-shaped proteins along DNA: case study of sliding clamps

Investigation of sliding DNA clamp dynamics by single-molecule fluorescence, mass spectrometry and structure-based modeling

Identification of β Clamp-DNA Interaction Regions That Impair the Ability of E. coli to Tolerate Specific Classes of DNA Damage

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