Review: The lord of the rings: Structure and mechanism of the sliding clamp loader

Kelch, Brian A.

doi:10.1002/bip.22827

Cited by 64 publications

(87 citation statements)

References 203 publications

(428 reference statements)

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“…It is also possible that Mcm5-Mcm3 ATP hydrolysis directly stimulates ring closure which causes Cdt1 release. Although ORC–Cdc6 has been proposed to function like a sliding-clamp loader during helicase loading 7 , of the known sliding clamp functions 26 , it appears this complex only retains the function of recruiting a protein-ring to the DNA. ORC and Cdc6 are not required to open the Mcm2–7 ring (Fig.…”

Section: Discussionmentioning

confidence: 99%

Mechanism and timing of Mcm2–7 ring closure during DNA replication origin licensing

Ticau

Friedman

Champasa

et al. 2017

Nat Struct Mol Biol

140

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Opening and closing of two ring-shaped Mcm2–7 DNA helicases is necessary to license eukaryotic origins of replication although the mechanisms controlling these events are unclear. The origin-recognition complex (ORC), Cdc6 and Cdt1 facilitate this process, establishing a topological link between each Mcm2–7 and origin DNA. Using colocalization single-molecule spectroscopy and single-molecule FRET (Förster resonance energy transfer), we monitored S. cerevisiae Mcm2–7 ring opening and closing during origin licensing. The two Mcm2–7 rings are open during initial DNA association and close sequentially, concomitant with release of their associated Cdt1. ATP hydrolysis by Mcm2–7 is coupled to ring closure and Cdt1 release, and failure to load the first Mcm2–7 prevents recruitment of the second Mcm2–7. Our findings identify key mechanisms controlling the Mcm2–7 DNA-entry gate during origin licensing and reveal that the two Mcm2–7 complexes are loaded by a coordinated series of events with implications for bidirectional replication initiation and quality control.

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Section: Discussionmentioning

confidence: 99%

Mechanism and timing of Mcm2–7 ring closure during DNA replication origin licensing

Ticau

Friedman

Champasa

et al. 2017

Nat Struct Mol Biol

140

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“…The eukaryotic DNA clamp proliferating-cell nuclear antigen (PCNA) is loaded onto the 3΄ end of a primer–template junction by replication factor C (RFC) and functions as a platform during DNA synthesis for loading of DNA polymerases and various factors involved in Okazaki-fragment processing, DNA-damage repair, chromatin assembly and sister-chromatid cohesion (1–4). RFC consists of one large subunit (RFC1) and four small subunits (RFC2–5), all of which belong to the AAA+ ATPase family (1,2,4).…”

Section: Introductionmentioning

confidence: 99%

“…RFC consists of one large subunit (RFC1) and four small subunits (RFC2–5), all of which belong to the AAA+ ATPase family (1,2,4). RFC hydrolyses ATP during PCNA loading.…”

Section: Introductionmentioning

confidence: 99%

Human CTF18-RFC clamp-loader complexed with non-synthesising DNA polymerase ε efficiently loads the PCNA sliding clamp

Fujisawa

Ohashi

Hirota

et al. 2017

Nucleic Acids Research

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The alternative proliferating-cell nuclear antigen (PCNA)-loader CTF18-RFC forms a stable complex with DNA polymerase ε (Polε). We observed that, under near-physiological conditions, CTF18-RFC alone loaded PCNA inefficiently, but loaded it efficiently when complexed with Polε. During efficient PCNA loading, CTF18-RFC and Polε assembled at a 3΄ primer–template junction cooperatively, and directed PCNA to the loading site. Site-specific photo-crosslinking of directly interacting proteins at the primer–template junction showed similar cooperative binding, in which the catalytic N-terminal portion of Polε acted as the major docking protein. In the PCNA-loading intermediate with ATPγS, binding of CTF18 to the DNA structures increased, suggesting transient access of CTF18-RFC to the primer terminus. Polε placed in DNA synthesis mode using a substrate DNA with a deoxidised 3΄ primer end did not stimulate PCNA loading, suggesting that DNA synthesis and PCNA loading are mutually exclusive at the 3΄ primer–template junction. Furthermore, PCNA and CTF18-RFC–Polε complex engaged in stable trimeric assembly on the template DNA and synthesised DNA efficiently. Thus, CTF18-RFC appears to be involved in leading-strand DNA synthesis through its interaction with Polε, and can load PCNA onto DNA when Polε is not in DNA synthesis mode to restore DNA synthesis.

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“…This follows in part because T4 is the simplest creature that utilizes the three major replication sub-assemblies characteristic of higher organisms, including a hexameric helicase–primase complex that ‘opens’ the duplex DNA genome, exposes the single-stranded (ss) DNA templates and primes (with short RNA sequences) DNA synthesis on the lagging strand template; a pair of DNA polymerases that engage in ‘coupled’ leading and lagging strand DNA synthesis (and initial ‘editing’ to assure high replication fidelity) within the ‘trombone-shaped’ DNA framework of the replication complex (7,8); and circular trimeric (or dimeric) replication clamps that are opened and closed and properly positioned on the polymerases by a pentameric complex that hydrolyses ATP and serves as a clamp-loader (and in many cases clamp-‘remover’) to control the processivity of DNA synthesis (9–11). …”

Section: Introductionmentioning

confidence: 99%

Single-molecule FRET studies of the cooperative and non-cooperative binding kinetics of the bacteriophage T4 single-stranded DNA binding protein (gp32) to ssDNA lattices at replication fork junctions

et al. 2016

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Gene 32 protein (gp32) is the single-stranded (ss) DNA binding protein of the bacteriophage T4. It binds transiently and cooperatively to ssDNA sequences exposed during the DNA replication process and regulates the interactions of the other sub-assemblies of the replication complex during the replication cycle. We here use single-molecule FRET techniques to build on previous thermodynamic studies of gp32 binding to initiate studies of the dynamics of the isolated and cooperative binding of gp32 molecules within the replication complex. DNA primer/template (p/t) constructs are used as models to determine the effects of ssDNA lattice length, gp32 concentration, salt concentration, binding cooperativity and binding polarity at p/t junctions. Hidden Markov models (HMMs) and transition density plots (TDPs) are used to characterize the dynamics of the multi-step assembly pathway of gp32 at p/t junctions of differing polarity, and show that isolated gp32 molecules bind to their ssDNA targets weakly and dissociate quickly, while cooperatively bound dimeric or trimeric clusters of gp32 bind much more tightly, can ‘slide’ on ssDNA sequences, and exhibit binding dynamics that depend on p/t junction polarities. The potential relationships of these binding dynamics to interactions with other components of the T4 DNA replication complex are discussed.

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Review: The lord of the rings: Structure and mechanism of the sliding clamp loader

Cited by 64 publications

References 203 publications

Mechanism and timing of Mcm2–7 ring closure during DNA replication origin licensing

Mechanism and timing of Mcm2–7 ring closure during DNA replication origin licensing

Human CTF18-RFC clamp-loader complexed with non-synthesising DNA polymerase ε efficiently loads the PCNA sliding clamp

Single-molecule FRET studies of the cooperative and non-cooperative binding kinetics of the bacteriophage T4 single-stranded DNA binding protein (gp32) to ssDNA lattices at replication fork junctions

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