The N-terminal domain of Escherichia coli RecA have multiple functions in promoting homologous recombination
© Lee and Wang; licensee BioMed Central Ltd. 2009
Received: 18 February 2009
Accepted: 01 April 2009
Published: 01 April 2009
Escherichia coli RecA mediates homologous recombination, a process essential to maintaining genome integrity. In the presence of ATP, RecA proteins bind a single-stranded DNA (ssDNA) to form a RecA-ssDNA presynaptic nucleoprotein filament that captures donor double-stranded DNA (dsDNA), searches for homology, and then catalyzes the strand exchange between ssDNA and dsDNA to produce a new heteroduplex DNA. Based upon a recently reported crystal structure of the RecA-ssDNA nucleoprotein filament, we carried out structural and functional studies of the N-terminal domain (NTD) of the RecA protein. The RecA NTD was thought to be required for monomer-monomer interaction. Here we report that it has two other distinct roles in promoting homologous recombination. It first facilitates the formation of a RecA-ssDNA presynaptic nucleoprotein filament by converting ATP to an ADP-Pi intermediate. Then, once the RecA-ssDNA presynaptic nucleoprotein filament is stably assembled in the presence of ATPγS, the NTD is required to capture donor dsDNA. Our results also suggest that the second function of NTD may be similar to that of Arg243 and Lys245, which were implicated earlier as binding sites of donor dsDNA. A two-step model is proposed to explain how a RecA-ssDNA presynaptic nucleoprotein filament interacts with donor dsDNA.
Escherichia coli RecA is the founding member of the RecA protein family. It is essential for the initiation of repair of DNA breaks via homologous recombination, induction of the DNA damage-induced 'SOS' response, and activation of translesion DNA synthesis, as well as development and transmission of antibiotic resistance genes [1, 2]. Nearly all known functions of RecA require the formation of a presynaptic helical filament comprised of single-stranded DNA (ssDNA) bound to multiple RecA monomers with ATP. During homologous recombination, this activated form of the helical filament is capable of interacting with homologous double-stranded DNA (dsDNA) to form a heteroduplex DNA molecule. Eventually, the DNA strands are exchanged, resulting in the displacement of one of the original duplex strands and the subsequent creation of a new heteroduplex (or D-loop). This function is evolutionarily conserved in other members of the RecA family, including archaeal RadA and the eukaryotic proteins, Rad51 and Dmc1.
The RecA monomer has three major structural domains: a small N-terminal domain (NTD), a core ATPase domain (CAD), and a large C-terminal domain (CTD). By contrast, the monomers of RadA/Rad51/Dmc1 consist of a CAD and a larger NTD. The CAD, often referred to as the RecA fold , is structurally similar to the ATPase domains of DNA/RNA helicases, F1 ATPases, chaperone-like ATPases, and membrane transporters . Highly conserved in all RecA family proteins, the CAD contains two disordered loops (denoted the L1 and L2 motifs) that bind to ssDNA and are responsible for ssDNA-stimulated ATPase activity . All RecA family proteins are polymerized via a polymerization motif located between the NTD and the CAD. The polymerization motif contains a hydrophobic residue (isoleucine 26 in E. coli RecA; phenylalanines in RadA, Rad51, and Dmc1) that docks within the hydrophobic pocket of the neighboring CAD. This interaction was also observed at the binding interface between a human Rad51 monomer and a BRC repeat of BRCA2 tumor suppressor protein [6–9]. The polymerization motif of archaeal RadA protein is responsible for the assembly of different quaternary structures, including toroidal rings, as well as right-handed and left-handed helical filaments [10, 11].
The crystal structures of RecA-ssDNA and RecA-dsDNA nucleoprotein complexes with ADP-AlF4--Mg2+ have recently been reported . These new structures have provided unprecedented new insights into the mechanisms and energetics of RecA protein. In the RecA-ssDNA-ADP-AlF4--Mg2+ filament complex, the ssDNA is bound by the L1 and L2 loop regions as well as by the N-terminal portion of the αF and αG helices that follow L1 and L2, respectively. The ADP-AlF4--Mg2+ is sandwiched between the CADs of two adjacent RecA protomers in a completely buried environment. AlF4- group is coordinated by the side chains of Lys248 and Lys250. Lys250 also hydrogen bonds to the side chain of Glu96 in the neighboring RecA protomer. Glu96 is the catalytic residue thought to activate a water molecule for nucleophilic attack on the γ-phosphate. This second interface is absent in the inactive filament, where the corresponding interface of the ATP analogue, adenylyl-imidodiphosphate (AMP-PNP), is solvent exposed. Therefore, the charged-stabilized hydrogen bonds that Lys248 and Lys250 make to the AlF4- group could explain the ATP-dependency of DNA binding, because the γ-phosphate of ATP is sensed across the RecA-RecA interface cooperating with DNA binding to promote the transition to the active filament state. Close to the filament axis, the ssDNA is extended 50% in length relative to B-form DNA with the same sequence. Each RecA monomer interacts with three nucleotides of the DNA (a triplet), and each triplet is also bound by three contiguous RecA monomers. Strikingly, the DNA-RecA interaction, or DNA extension, is not isotropic at the nucleotide level; instead, the DNA comprises a three nucleotide segment with a nearly normal B-form distance between bases (an axial rise of 3.5–4.2 Å for ssDNA and 3.2–4.5 Å for dsDNA), followed by a long, untwisted internucleotide stretch (~7.1–7.8 Å in ssDNA and 8.4 Å in ssDNA) before the next nucleotide triplet, in a repeating pattern. Such an unusual repeat pattern of DNA extension reveals a new structural basis of the dynamics of filament assembly in the presence of ssDNA, including initiation (or nucleation) of filament assembly and the observed cooperativity of RecA-DNA binding.
The RecA-dsDNA-ADP-AlF4--Mg2+ crystal structure was postulated to be an end product after the strand exchange reaction between a RecA-ssDNA nucleoprotein filament and a homologous dsDNA target [12, 13], implying that RecA protein filaments may complete all functions (including ssDNA binding, donor dsDNA capturing, and strand exchange) in right-handed forms and also within the filament axes. Here, we considered an alternative possibility: that the RecA-dsDNA crystal structures might simply represent annealing products of the ssDNA in the RecA-ssDNA nucleoprotein filament and a complementary ssDNA. First, in the RecA-ssDNA filament structure, the purine and pyrimidine bases of bound ssDNA are outwardly exposed. Second, the complementary ssDNA in the RecA-dsDNA structure makes very few physical contacts with the RecA protein filament, indicating that the annealing of these two ssDNAs has very little impact on the protein structures. Indeed, the overall protein structures of RecA-dsDNA filaments are highly similar to those of RecA-ssDNA-ADP-AlF4--Mg2+ structures [12, 13]. Because the molecular mechanism of the strand exchange reaction is still not understood, it is important to examine these two different possibilities further.
In the present study, we report that the NTD of RecA may have such a function. Additional mutant analysis revealed that it might have at least two distinct roles in promoting the RecA-mediated homologous recombination reaction.
Materials and methods
RecA protein production, enzymatic assays, and DNA substrates
An improved SUMO fusion protein expression system  was used to rapidly produce RecA proteins in E. coli. The nuclease assay, D-loop formation assay, ssDNA-dependent ATPase activity assay, and DNA substrates used in this study have also been described in detail . ATPγS (Adenosine 5'-O-(3-thio)triphosphate) and AMP-PNP were purchase from Sigma Aldrich.
The wild-type and mutant E. coli RecA proteins (2 μM) were incubated with 4 μM circular ΦX174 dsDNA (in base pairs [bps]) at 37°C for 30 min in reaction buffer (1 mM ATPγS, 10 mM Mg2+-acetate, 100 mM Na acetate, 25 mM Tris-Cl pH 7.4), and were chilled on ice to stop the reaction. A droplet (4 μl) was placed on a copper grid (300 mesh, Pelco, USA) coated with fresh carbon for 1 min at room temperature. Excess buffer was then carefully blotted away from the edge of the grid with Whatman #1 filter paper (Whatman Inc., USA). After staining for 4 min with 2.5% uranyl acetate, excess liquid was removed and the samples were air dried at room temperature. Bio-transmission EM was performed with a Tecnai G2 Spirit Bio TWIN (FEI Co., Netherlands) using an acceleration voltage of 120 kV. Images were recorded with a slowscan CCD camera (Gatan MultiScan 600) at a resolution of at least 1024 × 1024 pixels.
Duplex DNA capture assay
The duplex DNA capture assay was carried out using a protocol modified from that described previously for Rad51 and Hop2-Mnd1 proteins . To obtain presynaptic filaments for the dsDNA capture assay, a 5'-biotinylated ssDNA PA1656 (15 μM in nucleotides) and RecA proteins (5 or 20 μM) were mixed with 4 μL streptavidin-coated magnetic beads (Novagen, USA) in 20 μL of buffer C (20 mM HEPES-KOH at pH 7.0, 1 mM DTT, 100 ng/mL BSA, 2 mM Mg2+-acetate, 5% glycerol and 1 mM ATPγS) for 5 min at 37°C. BSA was used as a negative control for RecA-DNA binding reactions. The magnetic beads were isolated with a magnetic separator (Novagen). The supernatant (denoted as "S1") was set aside for later analysis. The magnetic beads were washed twice with 20 mL buffer C, and then mixed with 15 μM of donor dsDNA (bps) in 20 μL of buffer D (20 mM HEPES-KOH at pH 7.0, 1 mM DTT, 11 mM Mg2+-acetate, 5% glycerol) at 37°C for 10 min with gentle mixing every 1 min. This donor dsDNA was 300 bps in length, and its central region was homologous to PA1656. The magnetic beads were isolated again with a magnetic separator. The supernatant (denoted as "S2") was set aside for later analysis. After the magnetic beads were washed twice with 20 μL of buffer E (20 mM HEPES-KOH at pH 7.0, 1 mM DTT, 11 mM Mg2+-acetate, 5% glycerol and 1 mM ATPγS), proteins and DNA substrates were eluted by incubating with 20 mL of 1% SDS. The SDS eluates (denoted as "B") were separated on either a 12% SDS-PAGE stained with Coomassie blue to visualize bovine serum albumin (BSA) and RecA proteins, or a 1.5% agarose gel stained with ethidium bromide to visualize the donor dsDNA.
The NTD of RecA is similar in amino acid sequence to a functional motif of RadA/Rad51/Dmc1
The NTD of E. coli RecA contains only 33 amino acid residues and is much smaller in size than the RadA/Rad51/Dmc1 NTDs (> 60 amino acids). The crystal structure of the RecA-ssDNA presynaptic filament reveals that NTDs are located at the exterior surface of the helical filament. Moreover, NTDs are not directly involved in RecA-ssDNA interaction or RecA-ATP binding (Figure 1A) . A superimposition of RecA-RecA pairs from the active RecA-ssDNA presynaptic filament onto an inactive RecA protein filament (achieved by aligning the CADs) revealed a large conformational change in the hinge region (residues 31–40) that connects the NTD and CAD. As a result, these two filaments differ by a 32° rotation and an 18.5 Å translation of the CAD . This hinge region is located immediately after the polymerization motif (i.e., Ile26 of E. coli RecA), which functions as a fulcrum to mediate a large conformational change in response to ssDNA binding . A similar scenario was also reported in a structural transition of the RecA protein filament from a compressed form to a relaxed form . Intriguingly, RadA/Rad51/Dmc1 protein also contains a similar structural motif that we referred to previously as the subunit rotation motif (SRM) . The SRM also uses the polymerization motif as a fulcrum to mediate rotation along the central axis of the archaeal RadA protein polymer. Continuous clockwise axial rotation of archaeal RadA proteins is responsible for the progression of stepwise structural transitions: first from an inactive ring to a right-handed filament with 6 monomers per helical pitch, then to an overextended right-handed filament with 3 monomers per helical pitch, and, finally, to a left-handed filament with 4 monomers per helical pitch[10, 11, 18]. A key consequence of these structural transitions is the progressive relocation of the NTDs and the L1 ssDNA binding motif. The NTD has been shown to mediate donor dsDNA binding in both human Rad51  and archaeal RadA . In the RadA right-handed helical filament with 6 monomers per helical pitch, L1 resides near the axis filament and the NTD is located at the exterior surface of the filament. In contrast, in the overextended right-handed helical filament with 3 monomers per helical pitch, L1 relocates to the exterior surface of the filament and constitutes an outward-opening palm structure in combination with the NTD. Inside this palm structure, 5 conserved basic amino acid residues (Lys27 and Lys60 of the NTD; and Arg117, Arg223, and Arg229 of the L1 motif) of Sulfolobus solfataricus RadA (Sso RadA) surround this pocket (~25 Å in diameter), which may be wide enough to simultaneously accommodate an ssDNA and a donor dsDNA . All five positively charged residues are evolutionarily conserved in all archaeal and eukaryotic RecA family proteins. Therefore, the overextended right-handed filament structure of Sso RadA was proposed to represent a structural intermediate during the homologous search and pairing process of archaeal and eukaryotic RecA family proteins . Similarly, as described above, the RecA-ssDNA-ADP-AlF4--Mg2+ nucleoprotein filament was also overextended [12, 13].
Production of the wild type and mutant RecA proteins
Functional characterization of NTD mutant proteins
We then speculated that the wild-type, E18K, and E18K/K23A proteins might be better than other NTD mutant proteins at converting ATP into ADP-Pi. Accordingly, we compared the ATPase activities of these three RecA proteins in response to ssDNA. The ssDNA-dependent ATPase activity was determined as described before, by quantification of the inorganic phosphate (Pi) released from γ32P-labeled ATP . Indeed, wild-type, E18K, and E18K/K23A proteins all exhibited higher ssDNA-dependent ATPase activity than the other four NTD mutants (K23A, K23E, R33A, and R33E) (Figure 4D).
Taken together, the positively charged residues (Lys23, Arg33, or Lys18 created by the E18K mutation) at the outer surface of the NTD could facilitate the RecA conversion of ATP into intermediate ADP-Pi in response to ssDNA. This conversion could be avoided with the use of ATPγS, ATP-AlF4-, or even ADP-AlF4-. ATPγS is known to be better than ATP or AMP-PNP at stabilizing the RecA-ssDNA presynaptic nucleoprotein filament [1, 12, 24]. However, Lys23 and Arg33 apparently make no direct contact with ssDNA or ATP in the crystal structure of the RecA-ssDNA-ADP-AlF4--Mg2+ presynaptic nucleoprotein filament (Figure 1A) . It is likely that these positively charged residues facilitate a RecA protein filament to search for and capture ssDNA in the presence of ATP, and to then convert ATP into intermediate ADP-Pi. As a result, the protein filament binds ssDNA tightly to form an active presynaptic nucleoprotein filament.
Lys23 and Arg33 are indispensable for RecA's function in the presence of ATPγS
Lys23 and Arg33 are required for the RecA-ssDNA presynaptic nucleoprotein filament to capture dsDNA
Taken together, we conclude that both K23A/R33A and R243A/K245A mutants have two identical defects. First, both mutants had a lower binding affinity for ssDNA than the wild-type protein, because more mutant (20 μM) than wild-type protein (5 μM) was needed to assemble an equal amount of RecA-ssDNA presynaptic nucleoprotein filaments (Figure 7B). Second, once the RecA-ssDNA presynaptic filaments were assembled in the presence of ATPγS, both mutants were defective in capturing donor dsDNA (Figure 7C).
We report here that the E. coli RecA NTD exhibits limited but significant amino acid sequence homology with the NTD (specifically, the second H1hH2 motif) of the homologous proteins Rad51/Dmc1/RadA (Figure 2A). Notably, four basic residues (Lys8, Lys19, Lys23, and Arg33) of NTD constitute a positively charged helical patch along the outer surface of the RecA-ssDNA-ADP-AlF4--Mg2+ nucleoprotein filament. We also found that these four basic residues at the NTD have at least two distinct roles in promoting RecA-mediated homologous recombination by capturing DNA. First, they help the RecA protein, in response to ssDNA, to convert ATP into intermediate ADP-Pi, a process that can be avoided by use of ATPγS or ATP-AlF4-. Then, the NTD facilitates the RecA-ssDNA presynaptic nucleoprotein filament capture of donor dsDNA during the homologous search reaction. We think this function may be similar to those of the NTDs in eukaryotic and archaeal RecA proteins [18, 19].
The second function of the E. coli RecA NTD may be similar to that of Arg243 and Lys245. The NTD and Arg243/Lys245 might function at the same time to facilitate an active RecA-ssDNA-ATPγS nucleoprotein filament to capture dsDNA. However, judging from the crystal structure of the RecA-ssDNA-ADP-AlF4--Mg2+ nucleoprotein filament (Figure 1) , it is unlikely for a donor dsDNA to simultaneously contact both Lys23/Arg33 and Arg243/Lys245. This is because the basic amino acid residues of the NTD (including Lys8, Lys19, Lys23, and Arg33) are located along the outermost surface of the nucleoprotein filament, while Lys243 and Arg245 are embedded in the center of the nucleoprotein filament (Figure 1B). Therefore, we propose the following model to explain their functions during D-loop formation reaction. After an active RecA-ssDNA nucleoprotein filament is assembled in the presence of ATPγS (or ADP-Pi), the positively charged amino acid residues of the NTD will first non-specifically contact the phosphate groups of dsDNA, subsequently helping Lys243 and Arg245 to establish interactions with this donor dsDNA, and then carry out the strand exchange reaction. A prerequisite for this hypothesis is a rather large structural movement between the NTD and the CAD. This is not impossible. In fact, two similar incidents have been reported recently. First, during assembly of the RecA-ssDNA nucleoprotein filament, a hinge region that connects the NTD and CAD can undergo a large conformational change in response to ssDNA and ADP-AlF4--Mg2+. This hinge region is located immediately after the polymerization motif, which functions as a fulcrum to mediate this large conformational change . Second, we reported that clockwise axial rotation of the SRM of archaeal RadA proteins is responsible for a serial structural transition of RadA polymers from a protein ring to a right-handed helical filament, then to an over-wound right-handed helical filament, and finally to a left-handed helical filament. The SRM is located between the NTDs and CADs of RadA/Rad51/Dmc1 proteins, and it also uses the polymerization motif as a fulcrum to mediate this clockwise rotation along the helical filament axis [10, 11, 18]. It would be interesting to investigate whether the RecA NTD actually completes the rest of the dsDNA capture by rotating along the central axis of a presynaptic nucleoprotein filament.
Our results here are consistent with several previous studies on the RecA NTD. First, modeling of a 24-residue RecA N-terminal peptide revealed that the four positively charged residues (Lys6, Lys8, Lys19, and Lys23) were capable of binding DNA phosphate groups by electrostatic interactions . Second, it was reported that K6A/K19A and K6A/K23A double mutants both exhibited severe defects in RecA function in vivo . Third, deletion of the first N-terminal 9 amino acids of RecA (RecA-Δ9) manifested a rec- phenotype . The RecA-Δ9 protein loses both Lys6 and Lys8, although the authors emphasized only the lack of Lys6. They then postulated that the Δ9 mutant was defective in mediating essential monomer-monomer contact , because the ε-NH3 group of Lys6 forms a salt bridge with the carboxylate oxygen of the Asp139 side chain in the neighboring subunit . This hypothesis was correct for the free RecA protein filament, because the purified K6A mutant protein was indeed partly defective in oligomeric interaction in the absence of DNA . The Lys6-Asp139 salt bridge also exists in the RecA-ssDNA-ADP-AlF4--Mg2+ nucleoprotein filament . However, the K6A mutant protein demonstrated apparently normal formation of RecA-ssDNA helical filaments in vitro . In addition, the recA-K6A and rec A-K6D mutants both exhibited a rec+ phenotype . Therefore, a decrease in monomer-monomer contact due to breakage of the Lys6-Asp139 salt bridge is not likely to be the primary cause for the rec- phenotype of Δ9, K6A/K19A, and K6A/K23A mutants. Since the basic side chains of Lys8, Lys19, and Lys23 are located at the outer surface of RecA or RecA-ssDNA filaments to bind ssDNA or dsDNA, we think the rec- phenotype of Δ9, K6A/K19A, and K6A/K23A mutants may be the result of a loss or mutation of Lys8, Lys19, or Lys23, respectively.
In conclusion, we showed in this report that the NTD of RecA protein sequentially mediates ssDNA and dsDNA binding during homologous recombination. Our results also imply that a rather large rigid body movement between the NTD and CAD may be required for an active RecA-ssDNA-ATPγS-Mg2+nucleoprotein filament to carry out homology searching and binding to a target dsDNA. We suggest that such a rigid body movement is mediated by axial rotation of the NTD and CAD along the central axis of the RecA protein filament.
This work was supported by Academia Sinica (AS-97-FP-M02 to TFW) and the National Science Council, Taiwan (NSC96-2321-B-001-019 to TFW). We thank Dr. Yuan-Chih Chang (Institute of physics, Academia Sinica) for help in EM imaging.
- Cox MM: The bacterial RecA protein as a motor protein. Annu Rev Microbiol. 2003, 57: 551-577. 10.1146/annurev.micro.57.030502.090953.View ArticlePubMedGoogle Scholar
- Hastings PJ, Rosenberg SM, Slack A: Antibiotic-induced lateral transfer of antibiotic resistance. Trends Microbiol. 2004, 12: 401-404. 10.1016/j.tim.2004.07.003.View ArticlePubMedGoogle Scholar
- Story RM, Weber IT, Steitz TA: The structure of the E. coli RecA protein monomer and polymer. Nature. 1992, 355: 318-325. 10.1038/355318a0.View ArticlePubMedGoogle Scholar
- Wang JM: Nucleotide-dependent domain motions within rings of the RecA/AAA+ superfamily. J Struct Biol. 2004, 148: 259-267. 10.1016/j.jsb.2004.07.003.View ArticlePubMedGoogle Scholar
- Story RM, Steitz TA: Structure of the RecA protein-ADP complex. Nature. 1992, 355: 374-376. 10.1038/355374a0.View ArticlePubMedGoogle Scholar
- Chen PL, Chen CF, Chen Y, Xiao J, Sharp ZD, Lee WH: The BRC repeats in BRCA2 are critical for RAD51 binding and resistance to methyl methanesulfonate treatment. Proc Nat Acad Sci. 1998, 95: 5287-5292. 10.1073/pnas.95.9.5287.PubMed CentralView ArticlePubMedGoogle Scholar
- Yuan SS, Lee SY, Chen G, Song M, Tomlinson GE, Lee EY: BRCA2 is required for ionizing radiation-induced assembly of Rad51 complex in vivo. Cancer Res. 1999, 59: 3547-3551.PubMedGoogle Scholar
- Pellegrini L, Yu DS, Lo T, Anand S, Lee M, Blundell TL, Venkitaraman AR: Insights into DNA recombination from the structure of a RAD51-BRCA2 complex. Nature. 2002, 420: 287-293. 10.1038/nature01230.View ArticlePubMedGoogle Scholar
- Shin DS, Pellegrini L, Daniels DS, Yelent B, Craig L, Bates D, Yu DS, Shivji MK, Hitomi C, Arvai AS, Volkmann N, Tsuruta H, Blundell TL, Venkitaraman AR, Tainer JA: Full-length archaeal Rad51 structure and mutants: mechanisms for RAD51 assembly and control by BRCA2. EMBO J. 2003, 22: 4566-4576. 10.1093/emboj/cdg429.PubMed CentralView ArticlePubMedGoogle Scholar
- Chen LT, Ko TP, Chang YC, Lin KA, Chang CS, Wang AH, Wang TF: Crystal structure of the left-handed archaeal RadA helical filament: identification of a functional motif for controlling quaternary structures and enzymatic functions of RecA family proteins. Nucleic acids Res. 2007, 35: 1787-1801. 10.1093/nar/gkl1131.PubMed CentralView ArticlePubMedGoogle Scholar
- Wang TF, Chen LT, Wang AH: Right or left turn? RecA family protein filaments promote homologous recombination through clockwise axial rotation. BioEssays. 2008, 30: 48-56. 10.1002/bies.20694.View ArticlePubMedGoogle Scholar
- Chen Z, Yang H, Pavletich NP: Mechanism of homologous recombination from the RecA-ssDNA/dsDNA structures. Nature. 2008, 453: 489-484. 10.1038/nature06971.View ArticlePubMedGoogle Scholar
- Kowalczykowski SC: Structural biology: snapshots of DNA repair. Nature. 2008, 453: 463-466. 10.1038/453463a.View ArticlePubMedGoogle Scholar
- Rehrauer WM, Kowalczykowski SC: The DNA binding site(s) of the Escherichia coli RecA protein. J Biol Chem. 1996, 271: 11996-12002. 10.1074/jbc.271.39.23874.View ArticlePubMedGoogle Scholar
- Kurumizaka H, Ikawa S, Sarai A, Shibata T: The mutant RecA proteins, RecAR243Q and RecAK245N, exhibit defective DNA binding in homologous pairing. Arch Biochem Biophys. 1999, 365: 83-91. 10.1006/abbi.1999.1166.View ArticlePubMedGoogle Scholar
- Lee CD, Sun HC, Hu SM, Chiu CF, Homhuan A, Liang SM, Leng CH, Wang TF: An improved SUMO fusion protein system for effective production of native proteins. Protein Sci. 2008, 17: 1241-1248. 10.1110/ps.035188.108.PubMed CentralView ArticlePubMedGoogle Scholar
- Chi P, San Filippo J, Sehorn MG, Petukhova GV, Sung P: Bipartite stimulatory action of the Hop2-Mnd1 complex on the Rad51 recombinase. Genes Develop. 2007, 21: 1747-1757. 10.1101/gad.1563007.PubMed CentralView ArticlePubMedGoogle Scholar
- Chen LT, Ko TP, Chang YW, Lin KA, Wang AH, Wang TF: Structural and functional analyses of five conserved positively charged residues in the L1 and N-terminal DNA binding motifs of archaeal RADA protein. PLoS One. 2007, 2: e858-10.1371/journal.pone.0000858.PubMed CentralView ArticlePubMedGoogle Scholar
- Aihara H, Ito Y, Kurumizaka H, Yokoyama S, Shibata T: The N-terminal domain of the human Rad51 protein binds DNA: structure and a DNA binding surface as revealed by NMR. J Mol Biol. 1999, 290: 495-504. 10.1006/jmbi.1999.2904.View ArticlePubMedGoogle Scholar
- Karlin S, Brocchieri L: Evolutionary conservation of RecA genes in relation to protein structure and function. J Bacteriol. 1996, 178: 1881-1894.PubMed CentralPubMedGoogle Scholar
- McGrew DA, Knight KL: Molecular design and functional organization of the RecA protein. Crit Rev Biochem Mol Biol. 2003, 38: 385-432. 10.1080/10409230390242489.View ArticlePubMedGoogle Scholar
- Bi X, Corpina RA, Goldberg J: Structure of the Sec23/24-Sar1 pre-budding complex of the COPII vesicle coat. Nature. 2002, 419: 271-277. 10.1038/nature01040.View ArticlePubMedGoogle Scholar
- Bielli A, Haney CJ, Gabreski G, Watkins SC, Bannykh SI, Aridor M: Regulation of Sar1 NH2 terminus by GTP binding and hydrolysis promotes membrane deformation to control COPII vesicle fission. J Cell Biol. 2005, 171: 919-924. 10.1083/jcb.200509095.PubMed CentralView ArticlePubMedGoogle Scholar
- Joo C, McKinney SA, Nakamura M, Rasnik I, Myong S, Ha T: Real-time observation of RecA filament dynamics with single monomer resolution. Cell. 2006, 126: 515-527. 10.1016/j.cell.2006.06.042.View ArticlePubMedGoogle Scholar
- Kurumizaka H, Aihara H, Ikawa S, Kashima T, Bazemore LR, Kawasaki K, Sarai A, Radding CM, Shibata T: A possible role of the C-terminal domain of the RecA protein. A gateway model for double-stranded DNA binding. J Biol Chem. 1996, 271: 33515-33524. 10.1074/jbc.271.52.33515.View ArticlePubMedGoogle Scholar
- Aihara H, Ito Y, Kurumizaka H, Terada T, Yokoyama S, Shibata T: An interaction between a specified surface of the C-terminal domain of RecA protein and double-stranded DNA for homologous pairing. J Mol Biol. 1997, 274: 213-221. 10.1006/jmbi.1997.1403.View ArticlePubMedGoogle Scholar
- Zlotnick A, Brenner SL: An alpha-helical peptide model for electrostatic interactions of proteins with DNA. The N terminus of RecA. J Mol Biol. 1988, 209: 447-457. 10.1016/0022-2836(89)90009-0.View ArticleGoogle Scholar
- Morimatsu K, Horii T: Analysis of the DNA binding site of Escherichia coli RecA protein. Adv Biophys. 1995, 31: 23-48. 10.1016/0065-227X(95)99381-X.View ArticlePubMedGoogle Scholar
- Zaitsev EN, Kowalczykowski SC: Essential monomer-monomer contacts define the minimal length for the N-terminus of RecA protein. Mol Microbiol. 1998, 29: 1317-1318. 10.1046/j.1365-2958.1998.01006.x.View ArticlePubMedGoogle Scholar
- Eldin S, Forget AL, Lindenmuth DM, Logan KM, Knight KL: Mutations in the N-terminal region of RecA that disrupt the stability of free protein oligomers but not RecA-DNA complexes. J Mol Biol. 2000, 299: 91-101. 10.1006/jmbi.2000.3721.View ArticlePubMedGoogle Scholar
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