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The replication clamp-loading machine at work in the three domains of life

Nature Reviews Molecular Cell Biology volume 7, pages 751–761 (2006)Cite this article

Key Points

  • Processive DNA polymerases that replicate chromosomes interact with a ring-shaped clamp that encircles DNA and slides along the duplex.

  • All sliding clamps (Escherichia coli β-clamp, T4 bacteriophage gp45, archaeal and eukaryotic proliferating cell nuclear antigen (PCNA)) form similar planar ring structures with a central channel of sufficient width to encircle duplex DNA.

  • In all the clamps, the two faces of the ring are structurally distinct and polymerases and clamp loaders compete for binding to the same face. Therefore, the clamp loader must depart from the clamp for the DNA polymerase to function.

  • The sliding clamp is loaded onto DNA by a clamp-loader complex driven by ATP. Clamp loaders are circular heteropentameric complexes that have been conserved throughout evolution and their subunits are members of the AAA+ (ATPases associated with a variety of cellular activities) family of ATPases.

  • The ATP-binding sites of clamp loaders are located at subunit interfaces. Although clamp loaders are very similar in structure and function, the specifics of ATP use varies among different replication systems.

  • Insights into the way clamp loaders bind to DNA, recognize a primer–template junction, and open the clamp have been revealed by recent structural and biochemical studies. The findings of these studies indicate that the clamp assumes a right-handed spiral configuration when it opens, and it docks onto the helical surface of the clamp loader.

Abstract

Sliding clamps are ring-shaped proteins that tether DNA polymerases to DNA, which enables the rapid and processive synthesis of both leading and lagging strands at the replication fork. The clamp-loading machinery must repeatedly load sliding-clamp factors onto primed sites at the replication fork. Recent structural and biochemical analyses provide unique insights into how these clamp-loading ATPase machines function to load clamps onto the DNA. Moreover, these studies highlight the evolutionary conservation of the clamp-loading process in the three domains of life.

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Figure 1: Sliding clamps of different organisms.
Figure 2: The Escherichia coli γ clamp-loader complex.
Figure 3: The gp45 clamp and the gp44/62 clamp loader of bacteriophage T4.
Figure 4: The eukaryotic RFC–PCNA binary complex.
Figure 5: The archaeal RFC–PCNA–DNA ternary complex.

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Acknowledgements

We thank N. Yao and R. Georgescu for their artistic contributions to Figures 1 and 3. The authors are also grateful for a grant from the National Institutes of Health.

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  1. Rockefeller University, Howard Hughes Medical Institute, 1230 York Avenue, New York, 10021, New York, USA

    Chiara Indiani & Mike O'Donnell

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DATABASES

Protein Data Bank

1CZD

1GE8

1JR3

1JQL

1PLQ

1SXJ

2POL

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Glossary

Replicase

A DNA polymerase with accessory subunits, including a clamp and clamp loader, that replicates chromosomal DNA.

Protomers

Subunits from which a larger structure is built.

AAA+ family of ATPases

(ATPases associated with a variety of cellular activities). A large family of functionally divergent proteins that have a region of homology around the ATP sites. They are often used to remodel other macromolecules.

Primase

A specialized RNA polymerase that synthesizes short RNA primers to initiate DNA synthesis by DNA polymerases.

SRC motif

A three-residue motif (serine–arginine–cysteine) that is conserved in clamp-loading subunits and that contains a catalytic arginine residue that is involved in ATP hydrolysis.

FRET

(Fluorescence resonance energy transfer). The transfer of energy from a light-activated fluorophore to a second fluorophore that has a longer excitation wavelength. The efficiency of energy transfer depends on the distance between fluorophores.

Walker A and B motifs

These motifs are found in most ATP-binding proteins. The Walker A region, also known as a P-loop, binds ATP, whereas the Walker B region binds Mg2+. Both are required for ATP hydrolysis.

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Indiani, C., O'Donnell, M. The replication clamp-loading machine at work in the three domains of life. Nat Rev Mol Cell Biol 7, 751–761 (2006). https://doi.org/10.1038/nrm2022

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