Kuriyan Lab Logo        
Brian Kelch
       Kelch Stamp

Debora Makino
       Makino Stamp

How a DNA polymerase clamp loader opens a sliding clamp.

Kelch BA*, Makino DL*, O'Donnell M, Kuriyan J.

* These authors contributed equally

Science. 2011 Dec 23;334(6063):1675-80. (local copy)

Abstract / Movies / Figures from the paper / Supplemental Material / Coordinates / References


Processive chromosomal replication relies on sliding DNA clamps, which are loaded onto DNA by pentameric clamp loader complexes belonging to the AAA+ family of ATPases. We present structures for the ATP-bound state of the clamp loader complex from bacteriophage T4, bound to an open clamp and primer-template DNA. The clamp loader traps a spiral conformation of the open clamp so that both the loader and the clamp match the helical symmetry of DNA. One structure reveals that ATP has been hydrolyzed in one subunit, and suggests that clamp closure and ejection of the loader involves disruption of the ATP-dependent match in symmetry. The structures explain how synergy between the loader, the clamp and DNA can trigger ATP hydrolysis and release of the closed clamp on DNA.



Intro to Clamp Loader: Open Clamp: DNA Structure

Morphing Between Open and Closed States

Figures from the paper.

Click on the small image to get a bigger one.

First figure from paper

Figure 1. Clamp loaders and sliding clamps A) Clamp loading reaction. The clamp loader has low affinity for both clamp and primer-template DNA in the absence of ATP. Upon binding ATP, the clamp loader can bind the clamp and open it. The binding of primer-template DNA activates ATP hydrolysis, leading to ejection of the clamp loader. B) Three classes of clamp loaders. Bacterial clamp loaders are pentamers consisting of three proteins: δ (A position), γ (B, C and D), and δ' (E). Eukaryotic clamp loaders (RFC) consist of five different proteins, with the A subunit containing an A′ domain that bridges the gap between the A and E subunits. The T4 bacteriophage clamp loader consists of two proteins: gp44 (the B, C, D, & E subunits) and gp62 (the A subunit).
Illustrator file, local only

Second figure from paper

Figure 2. Architecture of the T4 clamp loader/clamp/DNA complex A) Structure of the T4 clamp loader bound to an open clamp. Ribbon and schematic diagrams of the complex between the T4 clamp loader (multi-colored), the open T4 clamp (gray), which is broken between subunits I and III, and primer-template DNA. The gp62 protein (the A subunit; red) bridges the gap in the clamp with its A domain (a vestigial AAA+ module) on the lower part of the clamp and its A′ domain at the top. The duplex region of primer-template DNA (orange) is bound in the interior of the clamp loader (yellow ribbon in the schematic indicates contacts from the clamp loader) and the central pore of the open clamp, with the template overhang extruded through the gap between the A and A′ domains. B) Interactions of the T4 clamp loader with the clamp. The six clamp interaction motifs of the clamp loader are displayed as surfaces with the remainder of the clamp loader shown as a thin ribbon. C) Two orthogonal views of a closed clamp bound to DNA and the T4 clamp loader as in crystal form I. The clamp interacts with the DNA phosphate backbone through arginine residues from each clamp subunit (yellow). D) Two orthogonal views of an open clamp bound to DNA and the T4 clamp loader. Diagram based on the open clamp complex in form II crystals. The sidechains of Arg 162 of subunit I and Arg 87 of subunit III are represented as sticks without the surface displayed. E) Two representations of distortions in the structure of the open clamp, relative to that of the closed clamp. Top, displacement vectors between the two structures are shown, scaled up by a factor of 4. The magnitude of the displacement is also indicated by color (blue to red). Vectors are drawn in the direction of displacement from the planar to open conformation and are derived from local alignments of each of the six pseudo-symmetric domains in the clamp trimer. Bottom, domain rotations derived from these local alignments are mapped onto a schematic diagram of the open clamp.
Illustrator file, local only

Third figure from paper

Figure 3. Symmetric and cooperative recognition of DNA. A) Spiral of AAA+ modules in the T4 clamp loader bound to an open clamp. The gp44 AAA+ modules, for which surfaces are displayed, form a spiral that tracks the minor groove of the DNA. The A subunit (gp62) is not shown. B) Arginine fingers and ATP coordinate the AAA+ spiral and DNA binding. A top-down view with the arginine fingers and the ATP analog (ADP•BeF3) shown as spheres (collar domains not shown). C) Cooperativity in ATP binding. The T4 clamp loader and the clamp, at concentrations of 2 and 5 μM, respectively, were incubated in the presence of 100 nM 5'TAMRA-labeled primer-template DNA. ADP•BeFx was titrated into the solution. As the concentration of the ATP analog increases, DNA binds to the clamp loader and the fluorescence anisotropy of the TAMRA probe increases in a highly sigmoidal fashion (napp ~ 3.3 ± 0.3).
PyMol files, local only

Fourth figure from paper

Figure 4. Hydrolysis-induced conformational changes. A) The fully ATP-bound conformation of the open clamp:clamp loader:DNA complex from form II crystals. The clamp loader holds the clamp such that there is a gap between subunits I and III of the clamp. B) Conformational changes in the clamp:clamp loader:DNA complex from ATP hydrolysis at the B subunit as seen in form III crystals. The structure of the complex is shown in the same orientation as in Fig. 4A (superposed using the AAA+ module of the C subunit). Clamp subunits I and III are sealed so that the clamp is now closed. The A and B subunits, as well as the entire collar region, undergo a large conformational change. C) Schematic diagram describing ATP hydrolysis-induced changes in clamp interactions. The clamp and the AAA+ modules of the clamp loader are shown from the side. In the ATP-loaded state, all AAA+ modules are positioned perfectly to match the clamp binding sites. Upon ATP hydrolysis, the B subunit swings away from the C subunit, altering the spacing between the clamp binding loops. D) ATP Hydrolysis severs contacts between AAA+ modules. The AAA+ modules are shown as surfaces. The interface between the B and C AAA+ modules has been completely disrupted.
Illustrator file, local only

Fifth figure from paper

Figure 5. A proposed DNA-dependent allosteric switch. A) The switch residue of the T4 clamp loader (Lys 80) is pointing into the interior chamber of the clamp loader to interact with DNA (DNA not shown). B) In the absence of DNA, the calculated electrostatic potential is extremely positive in the DNA binding cleft (calculation performed without Mg2+ or ADP•BeF3 ligands).
PyMol file, local only

Sixth figure from paper

Figure 6. A detailed mechanism for the clamp loading reaction. The reaction cycle for the T4 clamp loader is shown as a schematic diagram. (1) In the absence of ATP, the clamp loader AAA+ modules cannot organize into a spiral shape. (2) Upon ATP binding, the AAA+ modules form a spiral that can bind and open the clamp. (3) Primer-template DNA must thread through the gaps between the clamp subunits I & III and the clamp loader A and A′ domains. (4) Upon DNA binding in the interior chamber of the clamp loader, ATP hydrolysis is activated, most likely through flipping of the switch residue and release of the Walker B glutamate. (5) ATP hydrolysis at the B subunit breaks the interface at the AAA+ modules of the B and C subunits and allows closure of the clamp around primer-template DNA. Further ATP hydrolyses at the C and D subunits dissolve the symmetric spiral of AAA+ modules, thus ejecting the clamp loader because the recognition of DNA and the clamp is broken. The clamp is now loaded onto primer-template DNA and the clamp loader is free to recycle for another round of clamp loading.
Illustrator file, local only

Supplemental Material

Click on the small image to get a bigger one.

First supplimental figure
Fig. S1. Structural conservation of the A′ domain A) Positioning of the A′ domain in the yeast RFC/PCNA cocrystal structure(14). The A′ domain (boxed) is positioned against the E subunit and is lifted off of the clamp surface. Presumably, this domain would be contacting the upper side of the gap in the clamp when the clamp is opened. iig. S2. Fitting of EM reconstruction envelope from archaeal RFC/PCNA/DNA complex. B) Chain topology and orientation conservation. The positioning of the A′ domain in the yeast and T4 clamp loaders is shown. The insets are chain topology diagrams that show that the overall chain topology of the A′ domain of the yeast and T4 clamp loaders are shared. The overall orientation of the A′ domains relative to the E subunit is also conserved between yeast and T4 bacteriophage. (Due to the resolution of the yeast structure, many loops are missing and sequence register could not be identified in the electron density.)

Second supplimental figure
Figure S2: Fitting of EM reconstruction envelope from archaeal RFC/PCNA/DNA complex. A) Fitting of the T4 clamp loader:clamp:DNA crystal structure into the EM envelope. The structure from crystal form II was fit to the ~12 Å EM envelope of the P. furiosus clamp loader bound to DNA and an open form of PCNA(25). The fitting was performed in UCSF Chimera(60) to a volume calculated at 1σ. Most of the T4 structure fits remarkably well into the density, save for the A domain, which is degenerate in T4 but expected to be a AAA+ fold in archaeal clamp loaders(61). The A′ domain fits well into the density adjacent to the E subunit's ATPase site, illustrating that the A′ domain is conserved in loaders of trimeric clamps. Fig. S2. Fitting of EM reconstruction envelope from archaeal RFC/PCNA/DNA complex. B) Fitting of the E.coli clamp loader/DNA complex into the EM envelope. The structure of the E.coli clamp loader loaded with an ATP analog and primer-template DNA(15) was fit to the EM envelope as above. This structure matches the envelope quite well, save for the volume for the A′ domain, the clamp, and the duplex DNA extension, all of which were not present in the crystal structure.

Third supplimental figure
Figure S3: Model for entry of a primer template DNA into the clamp loader:clamp complex. Illustration of a possible mode of entry into the interior of the clamp loader for a primer-template junction. The major groove of the duplex at the primer-template junction or the ssDNA portion of the template can fit within the clamp opening. Given some flexibility in the dimensions of the open clamp, such a duplex or single-stranded DNA can slip through the opening and into the central chamber.

Fourth supplimental figure
Figure S4: Comparison of the open T4 clamp to molecular dynamics simulation of an open yeast clamp. Similarity between a yeast PCNA MD trajectory snapshot and the open clamp crystal structure. A snapshot (at 1ns) from the symmetrized MD simulation of yeast PCNA(52) was superposed onto the open clamp from crystal form II.

Fifth supplimental figure
Figure S5: Clamp Binding Interactions. A) Interactions of the A domain with the clamp. The A subunit projects a flexible tether deep into the clamp intrasubunit cleft with Leu3 and Phe4 providing most substantial interactions. This is the same cleft in the clamp to which the RB69 DNA polymerase binds(3) (RB69 polymerase peptide shown in yellow). In fact, the clamp binding motifs of gp62 and RB69 DNA polymerase share significant sequence homology (lower inset). B) A′ domain interactions. The A′ domain sits at the intersubunit cleft that is broken. Since half of the canonical binding site is across the gap in the clamp, the A′ domain exhibits the least interaction surface. C) gp44 binding into the intersubunit cleft. The interactions of the D subunit with the clamp are shown as an example (the B subunit binds in a similar manner into the symmetry-related cleft). This interface buries the second most surface area and is comprised mostly of Van der Waals interactions, as well as a salt bridge from Arg 162 of the clamp (gp45) to Asp 70 of gp44. D) gp44 binding into the intrasubunit cleft. The interactions of the E subunit with the clamp are shown as an example (the C subunit binds in a similar manner into the symmetry-related cleft). This interface is comprised almost exclusively of Van der Waals interactions.

Sixth supplimental figure
Figure S6: Interactions of the T4 clamp loader with DNA. A) Interactions of the AAA+ module with the template strand. Each AAA+ module binds 2.5 bp of duplex DNA. The D subunit's interactions are shown as a representative (the other subunits interact similarly). The amide nitrogen of Ile 81 is directly interacting with the phosphate group of Ade 16 which positions the positive end of the helix 4 dipole to stabilize the negative charge on the DNA backbone. The electron density is from an Fo-Fc sidechain omit map (all atoms past the Cβ omitted for highlighted residues) contoured at 2.7 σ and within 2.5 Å of the residues shown in sticks. B) Interactions of the A subunit with the ssDNA template overhang. The primer 3′ recessed end is bound under the collar domain, with the template single-stranded overhang extruded out of the central chamber and up along the collar domain. Only four nucleotides of the template overhang are actually visible in the electron density. The ssDNA is bound non-specifically by a series of hydrophobic stacking interactions between the bases (Phe 63 with Thy 11, Met 64 with Thy 10, Phe41 with both Thy 9 and Thy 8, and Tyr 108 with Thy 7). The phosphate groups of the nucleotides at the -1 and -2 positions are bound by Lys 113 and Lys 110, respectively. Omit density (contoured at 2.7 σ) for the single-stranded DNA template overhang is shown. C) The duplex DNA is distorted from B-form. The N-terminal helix dipole of helix 5 is positioned to extend into the minor groove. Ser 112 makes a hydrogen bond to the phosphate group on the primer backbone. D) B-form DNA is sterically prevented. Ideal B-form DNA was superposed using the phosphate groups of the template strand. The primer strand of ideal B-DNA clashes with the N-terminal end of helix 5, specifically Ser 112.

Seventh supplimental figure
Figure S7: Nucleotide state of the open clamp complex in crystal form II. A-D) Omit maps (nucleotide, magnesium and sidechain atoms past the Cβ omitted; contoured at 2.7σ are shown within 4Å of the omitted atoms) are shown for the ATP binding sites in the B, C, D and E sites (panels A, B, C, and D, respectively.) Density for ADP•BeF3 and Mg2+ is found in the B, C and D subunits, while the E subunit only has density for ADP and Mg2+.

Eighth supplimental figure
Figure S8: ATP binding cooperativity of clamp loaders. A) Cooperative ATP-binding of the E.coli clamp loader. DNA binding is used as readout for nucleotide binding. E.coli clamp loader (δδ′γ3) and β2 clamp (2μM and 5μM, respectively) were incubated in the presence of 100nM 5′TAMRA-labeled primer template DNA and ADP•BeF3 was titrated. As DNA becomes bound, TAMRA anisotropy increases. B) Primer-template DNA binding activity of the T4 clamp loader:clamp complex. The T4 clamp loader was titrated into a solution containing 1mM ADP•BeF3, 10μM clamp, and 100nM TAMRA-labeled primer-template DNA (42bp of duplex, 8nt of template strand overhang.) Fluorescence anisotropy of the TAMRA label is used as a readout for DNA binding. The T4 clamp loader binds with Kd of ~200nM.

First supplimental figure
Figure S9: Nucleotide state of the ternary complex with one subunit hydrolyzed (crystal form III). A-D) Omit maps (nucleotide, magnesium and sidechain atoms past the Cβ omitted; contoured at contoured at 2.7σ are shown within 4Å of the omitted atoms) are shown for the ATP binding sites in the B, C, D and E sites (panels A, B, C, and D, respectively.) The B subunit has density for only the ADP ligand and not the BeF3 moiety. Density for ADP•BeF3 and Mg2+ is found in the C and D subunits, while the E subunit only has density for Mg2+.

First supplimental figure
Figure S10: Hydrolysis in the T4 clamp loader induces similar motions in other AAA+ proteins. The T4 clamp loader, DnaA, and HslU AAA+ modules are shown in the same relative orientation in green, purple, and red, respectively. To illustrate the hydrolysis-induced domain motions, the ATP- and ADP-bound states of the three proteins are superposed using the N-terminal domain of the AAA+ module. The ATP-bound state for all proteins is shown in gray. Because the major motions can be described as rigid body rotation of domain II, only this domain of the ATP-loaded state is shown. While the degree of domain rotation is different in each AAA+ module (15°, 13°, and 5° for clamp loader, DnaA, and HslU, respectively), the direction of motion is conserved. The PDB codes for DnaA-ADP, DnaA-ATP, HslU-ADP, and HslU-ATP are from PDB codes: 1L8Q, 2HCB, 1G4A, and 1G3I, respectively.

First supplimental figure
Figure S11: A DNA-dependent allosteric switch for ATPase activity. A) Yeast clamp loader with the ATPase off. The Walker B catalytic glutamate (orange) is held in an inactive conformation by a conserved basic residue termed the switch residue (Arg 383 in yeast RFC-A; yellow) that interacts with the loop's carbonyl. B) T4 clamp loader with DNA and the ATPase on. DNA binding causes the switch (Lys 80 of gp44) to release the backbone of the Walker B glutamate and binds instead to the DNA phosphate backbone. The catalytic glutamate is now free to access the active conformation. C) The switch residue (Lys 100; yellow) in the E.coli B subunit is tucked into the interior of the AAA+ module in the absence of bound DNA(16). D) In presence of DNA, the switch residue is pointing out of the protein and makes ionic interactions with the phosphate backbone of DNA(15).


Coordinates in the Protein Data Bank:
Structure of T4 Bacteriophage clamp loader bound to the T4 clamp, primer-template DNA, and ATP analog
Structure of T4 Bacteriophage Clamp Loader Bound To Open Clamp, DNA and ATP Analog
Structure of T4 Bacteriophage Clamp Loader Bound To Closed Clamp, DNA and ATP Analog and ADP


1. Benkovic, S. J., Valentine, A. M. & Salinas, F. (2001). Replisome-mediated DNA replication. Annu Rev Biochem 70, 181-208.
2. Pomerantz, R. T. & O'Donnell, M. (2007). Replisome mechanics: insights into a twin DNA polymerase machine. Trends Microbiol 15, 156-64.
3. Shamoo, Y. & Steitz, T. A. (1999). Building a replisome from interacting pieces: sliding clamp complexed to a peptide from DNA polymerase and a polymerase editing complex. Cell 99, 155-66.
4. Krishna, T. S., Kong, X. P., Gary, S., Burgers, P. M. & Kuriyan, J. (1994). Crystal structure of the eukaryotic DNA polymerase processivity factor PCNA. Cell 79, 1233-43.
5. Kong, X. P., Onrust, R., O'Donnell, M. & Kuriyan, J. (1992). Three-dimensional structure of the beta subunit of E. coli DNA polymerase III holoenzyme: a sliding DNA clamp. Cell 69, 425-37.
6. Moarefi, I., Jeruzalmi, D., Turner, J., O'Donnell, M. & Kuriyan, J. (2000). Crystal structure of the DNA polymerase processivity factor of T4 bacteriophage. J Mol Biol 296, 1215-23.
7. Bailey, S., Wing, R. A. & Steitz, T. A. (2006). The structure of T. aquaticus DNA polymerase III is distinct from eukaryotic replicative DNA polymerases. Cell 126, 893-904.
8. Lamers, M. H., Georgescu, R. E., Lee, S.-G., O'Donnell, M. & Kuriyan, J. (2006). Crystal structure of the catalytic alpha subunit of E. coli replicative DNA polymerase III. Cell 126, 881-92.
9. Helt, C. E., Wang, W., Keng, P. C. & Bambara, R. A. (2005). Evidence that DNA damage detection machinery participates in DNA repair. Cell Cycle 4, 529-32.
10. Onrust, R., Stukenberg, P. T. & O'Donnell, M. (1991). Analysis of the ATPase subassembly which initiates processive DNA synthesis by DNA polymerase III holoenzyme. J Biol Chem 266, 21681-6.
11. Stukenberg, P. T., Studwell-Vaughan, P. S. & O'Donnell, M. (1991). Mechanism of the sliding beta-clamp of DNA polymerase III holoenzyme. J Biol Chem 266, 11328-34.
12. Neuwald, A. F., Aravind, L., Spouge, J. L. & Koonin, E. V. (1999). AAA+: A class of chaperone-like ATPases associated with the assembly, operation, and disassembly of protein complexes. Genome Res 9, 27-43.
13. Erzberger, J. P. & Berger, J. M. (2006). Evolutionary relationships and structural mechanisms of AAA+ proteins. Annu Rev Biophys Biomol Struct 35, 93-114.
14. Jeruzalmi, D., O'Donnell, M. & Kuriyan, J. (2001). Crystal structure of the processivity clamp loader gamma (gamma) complex of E. coli DNA polymerase III. Cell 106, 429-41.
15. Bowman, G. D., O'Donnell, M. & Kuriyan, J. (2004). Structural analysis of a eukaryotic sliding DNA clamp-clamp loader complex. Nature 429, 724-30.
16. Oyama, T., Ishino, Y., Cann, I. K., Ishino, S. & Morikawa, K. (2001). Atomic structure of the clamp loader small subunit from Pyrococcus furiosus. Mol Cell 8, 455-63.
17. Seybert, A., Singleton, M. R., Cook, N., Hall, D. R. & Wigley, D. B. (2006). Communication between subunits within an archaeal clamp-loader complex. EMBO J 25, 2209-18.
18. Guenther, B., Onrust, R., Sali, A., O'Donnell, M. & Kuriyan, J. (1997). Crystal structure of the delta' subunit of the clamp-loader complex of E. coli DNA polymerase III. Cell 91, 335-45.
19. Zhuang, Z., Berdis, A. J. & Benkovic, S. J. (2006). An alternative clamp loading pathway via the T4 clamp loader gp44/62-DNA complex. Biochemistry 45, 7976-89.
20. Hingorani, M. M. & O'Donnell, M. (1998). ATP binding to the Escherichia coli clamp loader powers opening of the ring-shaped clamp of DNA polymerase III holoenzyme. J Biol Chem 273, 24550-63.
21. Gomes, X. V. & Burgers, P. M. (2001). ATP utilization by yeast replication factor C. I. ATP-mediated interaction with DNA and with proliferating cell nuclear antigen. J Biol Chem 276, 34768-75.
22. Miyata, T., Suzuki, H., Oyama, T., Mayanagi, K., Ishino, Y. & Morikawa, K. (2005). Open clamp structure in the clamp-loading complex visualized by electron microscopic image analysis. Proc Natl Acad Sci USA 102, 13795-800.
23. Ason, B., Handayani, R., Williams, C. R., Bertram, J. G., Hingorani, M. M., O'Donnell, M., Goodman, M. F. & Bloom, L. B. (2003). Mechanism of loading the Escherichia coli DNA polymerase III beta sliding clamp on DNA. Bona fide primer/templates preferentially trigger the gamma complex to hydrolyze ATP and load the clamp. J Biol Chem 278, 10033-40.
24. Pietroni, P. & von Hippel, P. H. (2008). Multiple ATP binding is required to stabilize the "activated" (clamp open) clamp loader of the T4 DNA replication complex. J Biol Chem 283, 28338-53.
25. Simonetta, K. R., Kazmirski, S. L., Goedken, E. R., Cantor, A. J., Kelch, B. A., McNally, R., Seyedin, S. N., Makino, D. L., O'Donnell, M. & Kuriyan, J. (2009). The mechanism of ATP-dependent primer-template recognition by a clamp loader complex. Cell 137, 659-71.
26. Goedken, E. R., Kazmirski, S. L., Bowman, G. D., O'Donnell, M. & Kuriyan, J. (2005). Mapping the interaction of DNA with the Escherichia coli DNA polymerase clamp loader complex. Nat Struct Mol Biol 12, 183-90.
27. Turner, J., Hingorani, M. M., Kelman, Z. & O'Donnell, M. (1999). The internal workings of a DNA polymerase clamp-loading machine. EMBO J 18, 771-83.
28. Berdis, A. J. & Benkovic, S. J. (1996). Role of adenosine 5'-triphosphate hydrolysis in the assembly of the bacteriophage T4 DNA replication holoenzyme complex. Biochemistry 35, 9253-65.
29. Georgescu, R. E., Kim, S.-S., Yurieva, O., Kuriyan, J., Kong, X.-P. & O'Donnell, M. (2008). Structure of a sliding clamp on DNA. Cell 132, 43-54.
30. McNally, R., Bowman, G. D., Goedken, E. R., O'Donnell, M. & Kuriyan, J. (2010). Analysis of the role of PCNA-DNA contacts during clamp loading. BMC Struct Biol 10, 3.
31. Zhuang, Z., Yoder, B. L., Burgers, P. M. J. & Benkovic, S. J. (2006). The structure of a ring-opened proliferating cell nuclear antigen-replication factor C complex revealed by fluorescence energy transfer. Proc Natl Acad Sci USA 103, 2546-51.
32. Yao, N., Leu, F. P., Anjelkovic, J., Turner, J. & O'Donnell, M. (2000). DNA structure requirements for the Escherichia coli gamma complex clamp loader and DNA polymerase III holoenzyme. J Biol Chem 275, 11440-50.
33. Pluciennik, A., Dzantiev, L., Iyer, R. R., Constantin, N., Kadyrov, F. A. & Modrich, P. (2010). PCNA function in the activation and strand direction of MutL{alpha} endonuclease in mismatch repair. Proc Natl Acad Sci USA.
34. Nonin, S., Leroy, J. L. & Guéron, M. (1995). Terminal base pairs of oligodeoxynucleotides: imino proton exchange and fraying. Biochemistry 34, 10652-9.
35. Andreatta, D., Sen, S., Pérez Lustres, J. L., Kovalenko, S. A., Ernsting, N. P., Murphy, C. J., Coleman, R. S. & Berg, M. A. (2006). Ultrafast dynamics in DNA: "fraying" at the end of the helix. Journal of the American Chemical Society 128, 6885-92.
36. Kazmirski, S. L., Zhao, Y., Bowman, G. D., O'Donnell, M. & Kuriyan, J. (2005). Out-of-plane motions in open sliding clamps: molecular dynamics simulations of eukaryotic and archaeal proliferating cell nuclear antigen. Proc Natl Acad Sci USA 102, 13801-6.
37. Adelman, J. L., Chodera, J. D., Kuo, I.-F. W., Miller, T. F. & Barsky, D. (2010). The mechanical properties of PCNA: implications for the loading and function of a DNA sliding clamp. Biophys J 98, 3062-9.
38. Gulbis, J. M., Kelman, Z., Hurwitz, J., O'Donnell, M. & Kuriyan, J. (1996). Structure of the C-terminal region of p21(WAF1/CIP1) complexed with human PCNA. Cell 87, 297-306.
39. Yao, N., Turner, J., Kelman, Z., Stukenberg, P. T., Dean, F., Shechter, D., Pan, Z. Q., Hurwitz, J. & O'Donnell, M. (1996). Clamp loading, unloading and intrinsic stability of the PCNA, beta and gp45 sliding clamps of human, E. coli and T4 replicases. Genes Cells 1, 101-13.
40. Johnson, A. & O'Donnell, M. (2003). Ordered ATP hydrolysis in the gamma complex clamp loader AAA+ machine. J Biol Chem 278, 14406-13.
41. Ahmadian, M. R., Stege, P., Scheffzek, K. & Wittinghofer, A. (1997). Confirmation of the arginine-finger hypothesis for the GAP-stimulated GTP-hydrolysis reaction of Ras. Nat Struct Biol 4, 686-9.
42. Weiss, J. N. (1997). The Hill equation revisited: uses and misuses. FASEB J 11, 835-41.
43. Hinton, D. M. & Nossal, N. G. (1987). Bacteriophage T4 DNA primase-helicase. Characterization of oligomer synthesis by T4 61 protein alone and in conjunction with T4 41 protein. J Biol Chem 262, 10873-8.
44. Erzberger, J. P., Mott, M. L. & Berger, J. M. (2006). Structural basis for ATP-dependent DnaA assembly and replication-origin remodeling. Nat Struct Mol Biol 13, 676-83.
45. Wang, J., Song, J. J., Seong, I. S., Franklin, M. C., Kamtekar, S., Eom, S. H. & Chung, C. H. (2001). Nucleotide-dependent conformational changes in a protease-associated ATPase HsIU. Structure 9, 1107-16.
46. Gao, Y. Q., Yang, W. & Karplus, M. (2005). A structure-based model for the synthesis and hydrolysis of ATP by F1-ATPase. Cell 123, 195-205.
47. Abrahams, J. P., Leslie, A. G., Lutter, R. & Walker, J. E. (1994). Structure at 2.8 A resolution of F1-ATPase from bovine heart mitochondria. Nature 370, 621-8.
48. Seybert, A., Scott, D. J., Scaife, S., Singleton, M. R. & Wigley, D. B. (2002). Biochemical characterisation of the clamp/clamp loader proteins from the euryarchaeon Archaeoglobus fulgidus. Nucleic Acids Res 30, 4329-38.
49. Alley, S. C., Shier, V. K., Abel-Santos, E., Sexton, D. J., Soumillion, P. & Benkovic, S. J. (1999). Sliding clamp of the bacteriophage T4 polymerase has open and closed subunit interfaces in solution. Biochemistry 38, 7696-709.
50. Jeruzalmi, D., Yurieva, O., Zhao, Y., Young, M., Stewart, J., Hingorani, M., O'Donnell, M. & Kuriyan, J. (2001). Mechanism of processivity clamp opening by the delta subunit wrench of the clamp loader complex of E. coli DNA polymerase III. Cell 106, 417-28.
51. Enemark, E. J. & Joshua-Tor, L. (2006). Mechanism of DNA translocation in a replicative hexameric helicase. Nature 442, 270-5.
52. Glynn, S. E., Martin, A., Nager, A. R., Baker, T. A. & Sauer, R. T. (2009). Structures of asymmetric ClpX hexamers reveal nucleotide-dependent motions in a AAA+ protein-unfolding machine. Cell 139, 744-56.
53. Sancar, A. & Hearst, J. E. (1993). Molecular matchmakers. Science 259, 1415-20.
54. Young, M. C., Weitzel, S. E. & von Hippel, P. H. (1996). The kinetic mechanism of formation of the bacteriophage T4 DNA polymerase sliding clamp. J Mol Biol 264, 440-52.
55. Studier, F. W. (2005). Protein production by auto-induction in high density shaking cultures. Protein Expr Purif 41, 207-34.
56. Otwinowski, Z. M., W. (1997). Processing of X-ray Diffraction Data Collected in Oscillation Mode. In Methods in Enzymology (Carter, C. W. S., R.M., ed.), Vol. 276, pp. 307-326. Academic Press, New York.
57. McCoy, A. J., Grosse-Kunstleve, R. W., Adams, P. D., Winn, M. D., Storoni, L. C. & Read, R. J. (2007). Phaser crystallographic software. J Appl Crystallogr 40, 658-674.
58. Adams, P. D., Afonine, P. V., Bunkóczi, G., Chen, V. B., Davis, I. W., Echols, N., Headd, J. J., Hung, L.-W., Kapral, G. J., Grosse-Kunstleve, R. W., McCoy, A. J., Moriarty, N. W., Oeffner, R., Read, R. J., Richardson, D. C., Richardson, J. S., Terwilliger, T. C. & Zwart, P. H. (2010). PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr D Biol Crystallogr 66, 213-21.
59. Emsley, P. & Cowtan, K. (2004). Coot: model-building tools for molecular graphics. Acta Crystallogr D Biol Crystallogr 60, 2126-32.
60. Terwilliger, T. C. (2004). Using prime-and-switch phasing to reduce model bias in molecular replacement. Acta Crystallogr D Biol Crystallogr 60, 2144-9.
61. Pettersen, E. F., Goddard, T. D., Huang, C. C., Couch, G. S., Greenblatt, D. M., Meng, E. C. & Ferrin, T. E. (2004). UCSF Chimera--a visualization system for exploratory research and analysis. J Comput Chem 25, 1605-12.
62. Seybert, A. & Wigley, D. B. (2004). Distinct roles for ATP binding and hydrolysis at individual subunits of an archaeal clamp loader. EMBO J 23, 1360-71. Bowman, G.D., O'Donnell, M., and Kuriyan, J. (2004). Structural analysis of a eukaryotic sliding DNA clamp-clamp loader complex. Nature 429, 724-730.