Structural Mechanisms of DNA Replication, Repair, and Recombination

Melissa E. Stauffer and Walter J. Chazin

Departments of Biochemistry and Physics and Center for Structural Biology

Vanderbilt University, Nashville, TN 37232-8725 USA

DNA processing involves multiple biochemical steps requiring far more functions than

can be easily incorporated into a single protein. Consequently, replication, repair, and

recombination are performed by multi-protein assemblies. Remarkably, there is growing

evidence that these processes share many common features. The enzymes active in replicating

DNA during S phase are needed to synthesize new stretches of DNA during various types of

recombination and repair. Repair of all types must occur in tandem with replication and

transcription, since the exposure of DNA during these processes renders it open to discovery of

common lesions. Recombination is used to catalyze genetic crossover during meiosis, and is also

a major DNA repair mechanism during replicative and later phases of the cell cycle. Thus, the

processes of DNA replication, repair, and recombination are being increasingly viewed as

integrated events in cellular life.

Considerable progress has been made in developing techniques to investigate at the

atomic level how the DNA processing machinery functions. We are now in a position to ask very

basic questions: what are the structural mechanisms that regulate progression from initiation

through termination of a DNA processing pathway, and how does the DNA processing

machinery facilitate cross-talk between replication, recombination, and repair? We highlight here

some of the trends and concepts that are emerging as the structural mechanisms that regulate

DNA processing begin to be elucidated.

1

JBC Papers in Press. Published on April 16, 2004 as Manuscript R400015200

Copyright 2004 by The American Society for Biochemistry and Molecular Biology, Inc.

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

Multiple dynamic protein interactions coordinate the ordered progression of DNA

processing events. Working from a basic description of the hand-off of DNA from one protein to

another, we introduce the concepts of modularity of DNA processing proteins and the versatility

of the common folds found in these proteins. This is followed by recently reported examples of

direct competition and allosteric mechanisms promoting specific steps of DNA processing.

These concepts are integrated into a simple working model that provides a general framework for

thinking about the forward progression of DNA replication, repair, and recombination

assemblies.

The Concept of Handing Off

Evidence is mounting in favor of dynamic assembly and disassembly of the DNA

processing machinery at the time it is needed, and against the existence of preformed

holoenzyme complexes that meander around the cell until they are needed (1-3). Multiple

dynamic protein interactions are fundamental to the nature of DNA processing and to

progression through each processing pathway. The weak and/or transient nature of many of the

protein interactions is thought to be the basis for the ability of the DNA processing machinery to

assemble dynamically and to hand off the DNA from one subcomplex to the next. From the

perspective of the DNA, the proteins are sometimes described as ‘trading places’ (4).

A well-known example from replication is the competitive polymerase switching

observed during lagging strand synthesis (Figure 1A) (5). In eukaryotes, DNA polymerase α-

primase complex (pol-prim) synthesizes an RNA-DNA hybrid primer. For processive DNA

synthesis to follow, the sliding clamp, proliferating cell nuclear antigen (PCNA), must be loaded

by the clamp loader protein, replication factor C (RFC). Loading of PCNA triggers the

displacement of pol-prim from the 3’ end of the primer. Processive synthesis by DNA

polymerase δ (Pol δ) requires its direct association with PCNA. RFC is subsequently displaced

from PCNA, but remains stably associated with the PCNA/pol-prim complex during synthesis.

2

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

Each polymerase switch is facilitated by protein interactions with replication protein A (RPA), a

highly abundant ssDNA binding protein in eukaryotes (6).

Hand-off is also well documented in eukaryotic nucleotide excision repair (NER), during

which a wide variety of DNA lesions are removed and replaced (Figure 1B) (7). In the global

genome repair pathway, damage is detected by the XPC/hHR23B/centrin 2 heterotrimer. Local

unwinding is effected by recruited TFIIH, a heterodimer composed of XPB and XPD subunits.

Next, XPA and RPA associate with the complex and orient the incoming excision proteins,

ERCC1/XPF and XPG. Binding of XPG signals the release of XPC/hHR23B, while

ERCC1/XPF triggers the excision of damaged DNA and the release of XPA and TFIIH. Of these

proteins, only RPA remains stably bound after excision to facilitate association of the synthetic

factors RFC, PCNA, Pol δ, and DNA ligase.

Even in recombination, for which detailed knowledge lags somewhat behind that of

replication and repair, there are reports of proteins trading places on DNA (Figure 1C) (8,9).

Homologous recombination (HR) is one pathway used to repair double strand DNA breaks. In

eukaryotes, the 5’ ends around the break are resected by the Mre11/Rad50/NBS1 nuclease,

leaving 3’ ssDNA overhangs. These are coated by RPA, which binds with high affinity to

ssDNA. The displacement of RPA from ssDNA is effected by Rad51, the recombinase, with the

help of Rad52, a recombination mediator protein. Nucleation of Rad51 on ssDNA promotes

formation of the nucleoprotein filament that catalyzes the homology search and subsequent

strand exchange.

In each pathway, the sequential nature of the recruitment and dissociation of DNA

processing proteins is well established. The advantages of dynamic assembly and disassembly

are many, not the least of which is that fewer copies of ubiquitous proteins such as RPA and

PCNA are needed, since they can be recycled for use in other reactions. In addition, the ordered

hand-off of DNA from protein to protein (or complex to complex) allows for several levels of

temporal and spatial regulation of the processing pathways, while parallel steps between two (or

more) pathways allow for cross-talk.

3

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

Modular Proteins with Multiple Binding Sites

A common feature of proteins involved in DNA processing is modularity, the physical

connection of structural domains that perform different biochemical functions. This

characteristic facilitates the coordination of biochemical activities without constraining

functional modules to fit tightly together. Flexible tethering of domains in modular proteins

allows multiple steps to be carried out independently, yet remain coupled.

A corollary of modularity is the use of multiple contact points between one or more pairs

of interacting proteins. This facilitates the hand-off of processing proteins in situations where

rearrangement is required rather than full dissociation. It also provides a means to achieve a high

overall affinity from a set of weak individual interactions (vide infra). The use of multiple

binding sites also allows for more than one type of regulation at a single step of DNA processing.

During replication, the clamp loader protein RFC binds to RPA using three of its five

subunits (6). Evidence from replication studies suggests that there are two distinct binding sites

for RFC on RPA: one that overlaps with the Pol δ binding site and one that does not. Multiple

points of interaction between RFC and RPA facilitate hand-off of RFC without causing it to

completely dissociate. Pol δ competes with RFC for binding to RPA during the second

polymerase switch of lagging strand synthesis, but after Pol δ displaces RFC, RFC remains

bound to the polymerase holoenzyme complex. The maintenance of RFC in the holoenzyme is

mediated by a second, species-specific interaction with RPA (E. coli SSB cannot substitute) and

is essential for processive DNA synthesis.

During NER, XPA binds to both the 32-kDa and the 70-kDa subunit of RPA (RPA32 and

RPA70, respectively) (10,11). The Kd for the overall interaction measured by surface plasmon

resonance was 1.9 × 10-8 M (12). ELISA results suggested that the binding affinity is partitioned,

with 80% of the affinity assigned to RPA32 and 20% to RPA70 (11). The expected dissociation

constant for the XPA-RPA70 interaction was therefore on the order of 10-7 M. However, NMR

experiments used to characterize the interaction of the isolated domains revealed that their

4

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

binding affinity is several orders of magnitude weaker than the intact proteins’ (13). What factors

make the total affinity significantly stronger than the sum of the isolated domain interactions?

The key here is the linkage effect (14), which provides a means to obtain a high affinity

interaction between binding partners from the tethering together of two or more components that

by themselves interact very weakly. For example, the isolated A and B domains of RPA70 each

bind ssDNA with relatively modest affinity (Kd is micromolar). However, when tethered together

by a flexible ~10 residue linker, their overall affinity for ssDNA is nearly three orders of

magnitude higher (Kd is nanomolar) (15).

In recombination events, Rad52 has been shown to bind to both the A domain of RPA70

(RPA70A) and the C-terminal domain of RPA32 (RPA32C) (16). In this case, the different

points of contact between Rad52 and RPA are associated with different regulatory mechanisms.

NMR chemical shift perturbation assays demonstrated that residues 257-274 of Rad52 bind to

RPA32C at the same location as two other repair proteins: XPA and uracil DNA glycosylase

(17). Overlapping binding sites between proteins from three different repair pathways suggested

a competitive mechanism for switching between pathways. The interaction between Rad52 and

RPA32C is also responsible for an allosteric enhancement (5-fold) of the ssDNA binding affinity

of RPA, an effect mediated through the DNA binding domain in RPA32 (16). The same segment

of Rad52 (residues 218-303) binds to RPA70A, and sequence comparison between RPA32C and

RPA70A has predicted that the Rad52 binding site overlaps with the ssDNA binding site on

RPA70A. This interaction is expected to be competitive with both DNA and other proteins

known to bind at this site.

Versatile Protein Domains

Another striking observation about DNA processing proteins is the repeated utilization of

specific interaction modules (folded domains). Remarkably, these domains may interact

exclusively with proteins in one context, but will interact with both proteins and DNA in another,

and their sites of interaction may be similar, overlapping, or unique. Modules that interact with

5

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

both proteins and DNA are critical to the recruitment and positioning of peripheral processing

proteins as they assemble on and dissociate from the DNA.

The oligonucleotide/oligosaccharide-binding (OB) fold, the canonical ssDNA binding

domain (18,19), is one example of such a commonly used interaction module. OB-fold domains

are found in DNA processing proteins ranging from nucleases and pyrophosphatases to ssDNA-

and RNA-binding proteins (20). RPA has six OB folds (and one winged helix domain, see

below), four of which are involved in binding to ssDNA. In the crystal structure of the tandem

RPA70AB domains (residues 181-422 of RPA70), an oligonucleotide is bound in the continuous

cleft created by alignment of the tandem OB-fold domains (Figure 2A) (21). While OB-fold

domains are characterized and, in fact, named for binding to nucleotides, they are also involved

in protein interactions. The N-terminal domain of RPA70, for instance, is dedicated to

interactions with pol-prim and p53 (23), whereas the tandem RPA70AB domains are known to

interact with both ssDNA and proteins, including the SV40 large T antigen, XPA, and Rad51

(22).

A recent crystal structure of BRCA2, one of two well-known human breast cancer

associated proteins, reveals a fundamentally different mode of protein binding to an OB-fold

domain (23). BRCA2 possesses three OB-fold domains in its C-terminal region, two of which

are bound to a 9-mer nucleotide in the same manner as observed for RPA70AB (Figure 2B). The

third domain is bound to a fragment of a binding partner, DSS1, which is essential for

recombinational DNA repair (24). The DSS1 peptide binds away from the usual nucleotide

binding site into a positively charged groove that traverses the domain from front to back. Hence,

this OB-fold domain appears to have been adapted for a purpose other than nucleotide binding.

A second example of a versatile protein fold is the winged helix (WH) domain, a member

of the common helix-turn-helix superfamily (25). WH domains were first characterized in 1993,

and are best known for their role in dsDNA binding in transcription factors (26). Numerous

examples of DNA-bound structures are available for a wide functional variety of proteins. A

recently determined structure of the RecQ helicase from E. coli revealed that it contains a WH

6

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

domain that possesses the characteristic structural features of a DNA-binding WH domain (27).

Interestingly, a homologous portion of human Werner’s syndrome protein (WRN) is known to

binds proteins, such as Bloom’s syndrome protein and FEN-1, a eukaryotic endo/exonuclease

(28,29). The interaction of WRN with FEN-1 is functionally relevant, as it increases FEN-1’s

DNA cleavage activity 6-8 fold (28). Thus, the WH domain of WRN is either designed

specifically for protein interaction or it is bifunctional, with both protein and DNA binding

activity.

Insight into the structural basis distinguishing DNA and protein binding activities of

different WH domains can be obtained from comparative structural analysis. For example, the

RPA32C WH domain has no DNA binding activity and is strictly used for protein interaction

(17). The electrostatic surface of RPA32C contrasts greatly with that of the RecQ WH (Figure

2C). While the RecQ WH, which purportedly binds DNA, possesses a long tract of positive

charge along a shallow groove, RPA32C is mostly negative over the same surface, with no

observable groove for DNA binding.

A recent example of a dual-function WH domain is found in eukaryotic Cdc6, a

component of the pre-replication complex (30). Cdc6 has been shown to interact with the

minichromosome maintenance helicase in a way that inhibits helicase activity (31). The region of

Cdc6 responsible for the interaction is the conserved C-terminal WH domain. Interestingly, this

domain has also been demonstrated to bind dsDNA (32). The electrostatic surface (data not

shown) for this domain corroborates its dual functionality, since a combination of positive and

negative patches are found, and the positive surface corresponds to the canonical dsDNA binding

site of WH domains.

Competition for Binding Sites

As noted above, proteins in DNA processing assemblies usually have more than one

protein or DNA binding site. Often, the binding sites are clustered together or overlapping on a

single surface of a molecule, setting up a competitive mechanism for regulating progression

7

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

through a particular pathway. Direct competition between two or more molecules for a single

binding site facilitates hand-off of DNA and proteins during DNA processing. It allows for cell

cycle-coupled regulation via timed transcription of essential protein factors. It minimizes the

number of possible protein interactions, promoting interplay between alternate regulatory

mechanisms.

In DNA replication, competition is evident in the binding of bacterial polymerases I-V to

the β-clamp processivity factor (33). All the polymerases interact with the same hydrophobic

pocket into which the δ-subunit of the γ-complex clamp-loader binds. Competition for the

hydrophobic pocket of the β-clamp is therefore thought to provide the mechanism for switching

from the loading phase to the synthesis phase of replication. Also, the affinity of the β-clamp for

its partner proteins is modulated by its binding to DNA, and structural features of damaged DNA

may play a role in both successful bypass of lesions and coordination of different types of repair.

In eukaryotic replication, competition for RPA precludes simultaneous binding of pol-

prim, RFC, and Pol δ, promoting progression of replication from priming to synthesis. Each of

these proteins has a binding site on RPA, though the precise locations of binding are unknown

(6). The ability of proteins acting at later steps in the pathway to displace those present at earlier

steps suggests overlapping binding sites, which is a well-established feature of RPA. Mer et al.

showed that the C-terminal domain of the 32-kDa subunit of RPA binds to peptides from three

different repair proteins, all at the same site (17). This early finding suggested that competition

might facilitate handing off the DNA from one repair protein to another.

Two recent studies identify overlap between protein and ssDNA binding sites on RPA.

NMR chemical shift perturbation experiments have indicated that both XPA and Rad51 bind to

the canonical oligonucleotide binding site on RPA70A (13,34). NMR competition experiments

demonstrated that ssDNA could disrupt the preformed complex between RPA70A and the N-

terminal domain of Rad51 (Rad51N), indicating that the binding of Rad51N and ssDNA is

mutually exclusive (34). The competition between Rad51 and ssDNA for RPA is the basis for

8

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

the displacement of RPA during initiation of homologous recombination, while XPA affects the

preference of RPA for undamaged ssDNA during NER.

Homologous recombination is also regulated by competition in the case of the interaction

between BRCA2 and Rad51 (35). BRCA2 possesses eight BRC repeats in its central region, six

of which are capable of binding to Rad51 with varying affinities. The interaction is essential for

the transport of Rad51 into the nucleus and is inhibitory to Rad51 filament formation. The basis

for the inhibition is simply that the BRC sequence binds to the site on Rad51 at which it (Rad51)

self-associates. Thus, BRCA2 and Rad51 compete for an overlapping binding site on Rad51.

Still, Rad51 forms oligomeric filaments in vivo. How? One possible explanation is that the low-

affinity BRC repeats release Rad51 first, followed by release of the remaining tightly bound

molecules by allosteric or cooperative means.

Allostery

Allostery (action at a remote site) is a second common mechanism that promotes

progression through a DNA processing pathway. Allostery takes on many forms. Some protein

interactions serve to orient the binding partners for optimal activity at a distant site. In other

cases, ATP binding and/or hydrolysis are coupled to conformational changes that affect the ATP-

binding protein’s oligomerization state or its interactions with other proteins. Allostery can be

used in conjunction with other types of regulation to catalyze a single step of DNA processing.

Allostery is evident in the interactions between the eukaryotic sliding clamp protein,

PCNA, and its binding partners. PCNA is a recruitment factor for repair proteins from several

different pathways, including XPG (NER), uracil DNA glycosylase (base excision repair),

several MSH proteins (mismatch repair), and the Werner helicase (recombination) (36). The

homotrimeric structure of PCNA allows for several regulatory mechanisms, including the use of

multiple binding sites, competition for overlapping sites, and allosteric control of binding

partners. The type of DNA damage to which PCNA is bound plays a role in attracting binding

partners, several of which specifically recognize kinked or bent DNA (37,38). PCNA stimulates

9

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

the nuclease activity of flap endonuclease-1 (FEN-1), one of its binding partners for lagging

strand synthesis, by up to 50-fold (39). The interaction between these two proteins and its effect

on FEN-1’s DNA binding ability was recently characterized (40). The PCNA-FEN-1 interaction

induces the formation of a β-zipper, with strands contributed by the C-terminal segments of each

protein. On FEN-1, the β-zipper links the PCNA binding site to a DNA binding site. Formation

of the β-zipper likely enhances the nuclease activity of FEN-1 through allostery by properly

positioning it for cleavage of flapped DNA substrates.

Allosteric regulation is also observed for the Mre11/Rad50 complex, which generates the

3’ ssDNA ends used as templates for homologous recombination (41). Mre11 contains nuclease

activity specific for ssDNA, dsDNA ends, or hairpin DNA structures, while Rad50 plays a

structural role in bringing sister chromatids together. Recent crystal structures have elucidated

the allosteric mechanism whereby the ATPase activity of Rad50 is coupled to the nuclease

activity of Mre11 (42). Mre11 interacts with the ATPase domain of Rad50, which is comprised

of globular and coiled coil regions from two separate chains of the Rad50 homodimer. Catalytic

Rad50 heads are organized into a DNA-binding lobe (I) and a structural coiled coil lobe that

binds Mre11 (II). ATP binding to lobe I causes a 30° rotation of this lobe with respect to lobe II,

repositioning the bound DNA with respect to the Mre11 bound to lobe II (Figure 3). ATP

hydrolysis has the effect of increasing the kcat of nucleolysis by Mre11, which can be interpreted

primarily as an acceleration of product release in preparation for the next cleavage cycle.

During recombination, the binding and hydrolysis of ATP are coupled to strand exchange

through allosteric effects. ATP binding has recently been shown to affect the pitch and

orientation of subunits in the bacterial RecA filament (43). In eukaryotic Rad51, ATP hydrolysis

is coupled to strand exchange by the DNA-binding loop L1 adjacent to the ATP binding Walker

A motif (44). A mutation in this loop retained DNA-dependent ATPase activity, indicating that

primary DNA binding was not affected. However, subsequent measurements of strand exchange

showed no formation of joint molecules by the mutant protein, suggesting that ATP hydrolysis is

allosterically coupled to strand exchange via residues in loop L1.

10

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

Concluding Remarks

The accumulated evidence to date confirms that the DNA processing machinery is

dynamically assembled and disassembled in conjunction with the cell cycle and in response to

DNA damage. Assembly and rearrangement of protein assemblies is facilitated by the hand-off

of proteins and DNA from one step to the next. We have discussed five key concepts relevant to

understanding the progression of DNA processing assemblies:

1. Modular organization—DNA processing proteins contain multiple structural

domains with distinct biochemical functions.

2. Multiple contact points—DNA processing proteins interact with their partner

proteins through multiple sites.

3. Modest affinity—The majority of protein interactions characterized to date

have dissociation constants in the micromolar range.

4. Versatile structural modules—DNA processing proteins utilize a common set

of structural frameworks that can interact with DNA and/or protein.

5. Multiple structural mechanisms promote hand-off—Direct competition for

binding sites and allosteric structural rearrangements have been characterized.

The emerging view is that DNA processing assemblies are composed of modular proteins with

multiple, structurally independent functional domains. Individual contact points are characterized

by relatively low (micromolar) binding affinities, which promotes hand-off of DNA and proteins.

The structural modules commonly used by DNA processing proteins are versatile in their

binding preferences and modes. These characteristics are integrated into the structural

mechanisms that mediate the forward progression of DNA processing.

In this model, linked weak interactions provide high overall affinity while maintaining a

significant off-rate for each contact point (Figure 4).1 This in turn provides a ready means for

rapid disassembly: by simply blocking one of the weak interactions, the overall affinity between

two interacting molecules can be reduced by several orders of magnitude almost instantaneously.

11

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

The rapid transition between high and low affinity states promotes hand-off and forward

progression of DNA processing. This simple model provides a framework for developing a more

sophisticated and detailed understanding of the structural mechanisms used by the multi-protein

assemblies involved in DNA replication, repair, and recombination.

Figure Legends

Figure 1. Schematic diagrams of sequential hand-off of proteins during (A) replication, (B)

nucleotide excision repair, and (C) recombination. Proteins are labeled at the point at which

they become involved in each pathway.

Figure 2. Versatile proteins folds. OB-fold domains from (A) RPA70AB and (B) BRCA2. The

domains with ssDNA binding activity are colored yellow, with bound ssDNA in blue. The N-

terminal OB-fold from BRCA2 is colored orange, with the bound DSS1 peptide in red. The

black arrow indicates the location of the canonical ssDNA binding site. Panel C compares the

electrostatic surfaces of WH domains from the RecQ helicase (left) and RPA32C (right) in

equivalent orientations. Red and blue indicate negative and positive surface potential,

respectively. The arrows indicate the position of the canonical WH DNA binding helix 3.

Figure 3. Allosteric effect of ATP binding on the subdomain orientation of Rad50. Lobe I of

Rad50 (blue oval) contacts both DNA (black helix) and lobe II (purple oval). When ATP

binds (red cross), lobe I rotates with respect to lobe II, bringing the DNA into position for

nucleolytic cleavage by Mre11 (yellow diamond) bound to lobe II.

Figure 4. Key concepts in DNA processing. The upper panel symbolizes the interaction between

two multidomain proteins (A, two open rectangles; B, two filled ovals) that have two contact

points. The overall affinity between the two proteins results from two modest affinity

interactions, each of which has an appreciable off-rate. This corresponds to one

representation of the linkage effect. The lower panel represents the facilitation of hand-off in

this system. A third protein (checked circle) needs to bind to only one of the two domains of

12

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

protein B in order to drastically reduce the overall binding affinity and promote release of B

from A.

References

1. Kowalczykowski, S. C. (2000) Nat Struct Biol 7, 1087-1089

2. Rademakers, S., Volker, M., Hoogstraten, D., Nigg, A. L., Mone, M. J., Van Zeeland, A.

A., Hoeijmakers, J. H., Houtsmuller, A. B., and Vermeulen, W. (2003) Mol Cell Biol 23,

5755-5767

3. Essers, J., Houtsmuller, A. B., van Veelen, L., Paulusma, C., Nigg, A. L., Pastink, A.,

Vermeulen, W., Hoeijmakers, J. H., and Kanaar, R. (2002) Embo J 21, 2030-2037

4. Yuzhakov, A., Kelman, Z., and O'Donnell, M. (1999) Cell 96, 153-163

5. Davey, M. J., and O'Donnell, M. (2000) Curr Opin Chem Biol 4, 581-586

6. Yuzhakov, A., Kelman, Z., Hurwitz, J., and O'Donnell, M. (1999) Embo J 18, 6189-6199

7. Riedl, T., Hanaoka, F., and Egly, J. M. (2003) Embo J 22, 5293-5303

8. Sung, P., Krejci, L., Van Komen, S., and Sehorn, M. G. (2003) J Biol Chem 278, 42729-

42732

9. Valerie, K., and Povirk, L. F. (2003) Oncogene 22, 5792-5812

10. Stigger, E., Drissi, R., and Lee, S. H. (1998) J Biol Chem 273, 9337-9343

11. Walther, A. P., Gomes, X. V., Lao, Y., Lee, C. G., and Wold, M. S. (1999) Biochemistry

38, 3963-3973

12. Saijo, M., Kuraoka, I., Masutani, C., Hanaoka, F., and Tanaka, K. (1996) Nucleic Acids

Res 24, 4719-4724

13. Daughdrill, G. W., Buchko, G. W., Botuyan, M. V., Arrowsmith, C., Wold, M. S.,

Kennedy, M. A., and Lowry, D. F. (2003) Nucleic Acids Res 31, 4176-4183

14. Zhou, H. X. (2001) Biochemistry 40, 15069-15073

15. Arunkumar, A. I., Stauffer, M. E., Bochkareva, E., Bochkarev, A., and Chazin, W. J.

(2003) J Biol Chem 278, 41077-41082

13

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

16. Jackson, D., Dhar, K., Wahl, J. K., Wold, M. S., and Borgstahl, G. E. (2002) J Mol Biol

321, 133-148

17. Mer, G., Bochkarev, A., Gupta, R., Bochkareva, E., Frappier, L., Ingles, C. J., Edwards,

A. M., and Chazin, W. J. (2000) Cell 103, 449-456

18. Arcus, V. (2002) Curr Opin Struct Biol 12, 794-801

19. Bochkarev, A., and Bochkareva, E. (2004) Curr Opin Struct Biol 14, 36-42

20. Theobald, D. L., Mitton-Fry, R. M., and Wuttke, D. S. (2003) Annu Rev Biophys Biomol

Struct 32, 115-133

21. Bochkarev, A., Pfuetzner, R. A., Edwards, A. M., and Frappier, L. (1997) Nature 385,

176-181

22. Iftode, C., Daniely, Y., and Borowiec, J. A. (1999) Crit Rev Biochem Mol Biol 34, 141-

180

23. Yang, H., Jeffrey, P. D., Miller, J., Kinnucan, E., Sun, Y., Thoma, N. H., Zheng, N.,

Chen, P. L., Lee, W. H., and Pavletich, N. P. (2002) Science 297, 1837-1848

24. Kojic, M., Yang, H., Kostrub, C. F., Pavletich, N. P., and Holloman, W. K. (2003) Mol

Cell 12, 1043-1049

25. Gajiwala, K. S., and Burley, S. K. (2000) Curr Opin Struct Biol 10, 110-116

26. Giraldo, R. (2003) FEMS Microbiol Rev 26, 533-554

27. Bernstein, D. A., Zittel, M. C., and Keck, J. L. (2003) Embo J 22, 4910-4921

28. Brosh, R. M., Jr., von Kobbe, C., Sommers, J. A., Karmakar, P., Opresko, P. L.,

Piotrowski, J., Dianova, I., Dianov, G. L., and Bohr, V. A. (2001) Embo J 20, 5791-5801

29. von Kobbe, C., Karmakar, P., Dawut, L., Opresko, P., Zeng, X., Brosh, R. M., Jr.,

Hickson, I. D., and Bohr, V. A. (2002) J Biol Chem 277, 22035-22044

30. Liu, J., Smith, C. L., DeRyckere, D., DeAngelis, K., Martin, G. S., and Berger, J. M.

(2000) Mol Cell 6, 637-648

31. Shin, J. H., Grabowski, B., Kasiviswanathan, R., Bell, S. D., and Kelman, Z. (2003) J

Biol Chem 278, 38059-38067

14

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

32. Grabowski, B., and Kelman, Z. (2001) J Bacteriol 183, 5459-5464

33. Lopez de Saro, F. J., Georgescu, R. E., Goodman, M. F., and O'Donnell, M. (2003) Embo

J 22, 6408-6418

34. Stauffer, M. E., and Chazin, W. J. (2004) Journal of Biological Chemistry, in press

35. Pellegrini, L., Yu, D. S., Lo, T., Anand, S., Lee, M., Blundell, T. L., and Venkitaraman,

A. R. (2002) Nature 420, 287-293

36. Maga, G., and Hubscher, U. (2003) J Cell Sci 116, 3051-3060

37. Sawaya, M. R., Prasad, R., Wilson, S. H., Kraut, J., and Pelletier, H. (1997) Biochemistry

36, 11205-11215

38. Lee, J. Y., Chang, C., Song, H. K., Moon, J., Yang, J. K., Kim, H. K., Kwon, S. T., and

Suh, S. W. (2000) Embo J 19, 1119-1129

39. Tom, S., Henricksen, L. A., and Bambara, R. A. (2000) J Biol Chem 275, 10498-10505

40. Chapados, B. R., Hosfield, D. J., Han, S., Qiu, J., Yelent, B., Shen, B., and Tainer, J. A.

(2004) Cell 116, 39-50

41. Hopfner, K. P., and Tainer, J. A. (2003) Curr Opin Struct Biol 13, 249-255

42. Hopfner, K. P., Karcher, A., Craig, L., Woo, T. T., Carney, J. P., and Tainer, J. A. (2001)

Cell 105, 473-485

43. VanLoock, M. S., Yu, X., Yang, S., Lai, A. L., Low, C., Campbell, M. J., and Egelman,

E. H. (2003) Structure (Camb) 11, 187-196

44. Shin, D. S., Pellegrini, L., Daniels, D. S., Yelent, B., Craig, L., Bates, D., Yu, D. S.,

Shivji, M. K., Hitomi, C., Arvai, A. S., Volkmann, N., Tsuruta, H., Blundell, T. L.,

Venkitaraman, A. R., and Tainer, J. A. (2003) Embo J 22, 4566-4576

Footnotes 1 We note here there is mixing of the kinetic and thermodynamic views, but the assumption that

weak binding is associated with high on/off rates is logical in this context.

15

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

R P A

P C NA R FC αA

B

XP C

T F IIH R P A

XP A

XP F XP G

CR P A

R ad52R ad51

δ

δ

α

F igure 1

16

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

A

B

C

F igure 2

A

B

C

17

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

50-I50-II

50-I

ATP

11

F igure 3

50-II11

30o

18

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

F igure 4

19

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from

Melissa E. Stauffer and Walter J. ChazinStructural mechanisms of DNA replication, repair, and recombination

published online April 16, 2004J. Biol. Chem. 

  10.1074/jbc.R400015200Access the most updated version of this article at doi:

 Alerts:

  When a correction for this article is posted• 

When this article is cited• 

to choose from all of JBC's e-mail alertsClick here

by guest on June 11, 2018http://w

ww

.jbc.org/D

ownloaded from