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Three new structures of left-handed RADA helical filaments: structural flexibility of N-terminal domain is critical for recombinase activity.

Chang YW, Ko TP, Lee CD, Chang YC, Lin KA, Chang CS, Wang AH, Wang TF - PLoS ONE (2009)

Bottom Line: We recently reported a structure of RadA left-handed helical filament, and here present three new structures of RadA left-handed helical filaments.Mutant analysis further confirms that structural flexibility of NTD is essential for RadA's recombinase activity.These results support our previous hypothesis that ATP-dependent axial rotation of RadA nucleoprotein helical filament promotes homologous recombination.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemical Science, National Taiwan University, Taipei, Taiwan.

ABSTRACT
RecA family proteins, including bacterial RecA, archaeal RadA, and eukaryotic Dmc1 and Rad51, mediate homologous recombination, a reaction essential for maintaining genome integrity. In the presence of ATP, these proteins bind a single-strand DNA to form a right-handed nucleoprotein filament, which catalyzes pairing and strand exchange with a homologous double-stranded DNA (dsDNA), by as-yet unknown mechanisms. We recently reported a structure of RadA left-handed helical filament, and here present three new structures of RadA left-handed helical filaments. Comparative structural analysis between different RadA/Rad51 helical filaments reveals that the N-terminal domain (NTD) of RadA/Rad51, implicated in dsDNA binding, is highly flexible. We identify a hinge region between NTD and polymerization motif as responsible for rigid body movement of NTD. Mutant analysis further confirms that structural flexibility of NTD is essential for RadA's recombinase activity. These results support our previous hypothesis that ATP-dependent axial rotation of RadA nucleoprotein helical filament promotes homologous recombination.

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Crystal structures of three new left-handed RadA helical filaments.The ribbon representations of RadA dimers (A) and helical filaments (B). Each RadA promotor is indicated by different colors. The helical pitches and protein database accession codes of these three helical filaments are also indicated, respectively.
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pone-0004890-g001: Crystal structures of three new left-handed RadA helical filaments.The ribbon representations of RadA dimers (A) and helical filaments (B). Each RadA promotor is indicated by different colors. The helical pitches and protein database accession codes of these three helical filaments are also indicated, respectively.

Mentions: The first left-handed helical filament (2DFL) that we reported [22] has a helical pitch of 125.6 Å and contains 4 identical monomers per helical turn. Here we report additional three new crystal structures of left-handed helical filaments. Their helical pitches are 130.4 Å (2ZUB), 136.1 Å (2ZUC) and 132.8 Å (2ZUD), respectively (Figure 1). These crystals all belong to the space group P212121 (Table 1). Unlike 2DFL, these three new left-handed helical filament structures are composed of two identical RadA dimers in each helical turn, and the two protomers in each dimer are structurally different. It was reported before that RecA family proteins might function as a dimer. First, the helical filament structure (PDB accession code 1SZP) of the yeast Rad51-I345T gain-of-function mutant suggested that the functional unit of Rad51 might be a dimer [16]. Second, a study of the EcRecA fused dimer also indicated that dimeric RecA might be the functional unit for assembly of active nucleoprotein filaments and for their coordinated activities during homologous recombination [24]. Therefore, a dimeric functional unit could exist not only in right-handed filaments but also in left-handed helical filaments.


Three new structures of left-handed RADA helical filaments: structural flexibility of N-terminal domain is critical for recombinase activity.

Chang YW, Ko TP, Lee CD, Chang YC, Lin KA, Chang CS, Wang AH, Wang TF - PLoS ONE (2009)

Crystal structures of three new left-handed RadA helical filaments.The ribbon representations of RadA dimers (A) and helical filaments (B). Each RadA promotor is indicated by different colors. The helical pitches and protein database accession codes of these three helical filaments are also indicated, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2654063&req=5

pone-0004890-g001: Crystal structures of three new left-handed RadA helical filaments.The ribbon representations of RadA dimers (A) and helical filaments (B). Each RadA promotor is indicated by different colors. The helical pitches and protein database accession codes of these three helical filaments are also indicated, respectively.
Mentions: The first left-handed helical filament (2DFL) that we reported [22] has a helical pitch of 125.6 Å and contains 4 identical monomers per helical turn. Here we report additional three new crystal structures of left-handed helical filaments. Their helical pitches are 130.4 Å (2ZUB), 136.1 Å (2ZUC) and 132.8 Å (2ZUD), respectively (Figure 1). These crystals all belong to the space group P212121 (Table 1). Unlike 2DFL, these three new left-handed helical filament structures are composed of two identical RadA dimers in each helical turn, and the two protomers in each dimer are structurally different. It was reported before that RecA family proteins might function as a dimer. First, the helical filament structure (PDB accession code 1SZP) of the yeast Rad51-I345T gain-of-function mutant suggested that the functional unit of Rad51 might be a dimer [16]. Second, a study of the EcRecA fused dimer also indicated that dimeric RecA might be the functional unit for assembly of active nucleoprotein filaments and for their coordinated activities during homologous recombination [24]. Therefore, a dimeric functional unit could exist not only in right-handed filaments but also in left-handed helical filaments.

Bottom Line: We recently reported a structure of RadA left-handed helical filament, and here present three new structures of RadA left-handed helical filaments.Mutant analysis further confirms that structural flexibility of NTD is essential for RadA's recombinase activity.These results support our previous hypothesis that ATP-dependent axial rotation of RadA nucleoprotein helical filament promotes homologous recombination.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemical Science, National Taiwan University, Taipei, Taiwan.

ABSTRACT
RecA family proteins, including bacterial RecA, archaeal RadA, and eukaryotic Dmc1 and Rad51, mediate homologous recombination, a reaction essential for maintaining genome integrity. In the presence of ATP, these proteins bind a single-strand DNA to form a right-handed nucleoprotein filament, which catalyzes pairing and strand exchange with a homologous double-stranded DNA (dsDNA), by as-yet unknown mechanisms. We recently reported a structure of RadA left-handed helical filament, and here present three new structures of RadA left-handed helical filaments. Comparative structural analysis between different RadA/Rad51 helical filaments reveals that the N-terminal domain (NTD) of RadA/Rad51, implicated in dsDNA binding, is highly flexible. We identify a hinge region between NTD and polymerization motif as responsible for rigid body movement of NTD. Mutant analysis further confirms that structural flexibility of NTD is essential for RadA's recombinase activity. These results support our previous hypothesis that ATP-dependent axial rotation of RadA nucleoprotein helical filament promotes homologous recombination.

Show MeSH
Related in: MedlinePlus