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Crystal structure of the Holliday junction migration motor protein RuvB from Thermus thermophilus HB8 - PubMed

  • ️Mon Jan 01 2001

Crystal structure of the Holliday junction migration motor protein RuvB from Thermus thermophilus HB8

K Yamada et al. Proc Natl Acad Sci U S A. 2001.

Abstract

We report here the crystal structure of the RuvB motor protein from Thermus thermophilus HB8, which drives branch migration of the Holliday junction during homologous recombination. RuvB has a crescent-like architecture consisting of three consecutive domains, the first two of which are involved in ATP binding and hydrolysis. DNA is likely to interact with a large basic cleft, which encompasses the ATP-binding pocket and domain boundaries, whereas the junction-recognition protein RuvA may bind a flexible beta-hairpin protruding from the N-terminal domain. The structures of two subunits, related by a noncrystallographic pseudo-2-fold axis, imply that conformational changes of motor protein coupled with ATP hydrolysis may reflect motility essential for its translocation around double-stranded DNA.

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Figures

Figure 1
Figure 1

Structure and topology of RuvB from Tth. (a) Experimental (Left) and final 2Fo − Fc (Right) electron density maps determined for the central β-sheet region of domain N. (b) Stereoview of the overall fold bound to a AMPPNP. The monomer is viewed from the nucleotide-binding side. Domains N, M, and C are colored in blue, yellow, and green, respectively. Walker motifs and sensor motifs are colored in magenta and orange, respectively. Residues involved in RuvB activities are labeled by green (RuvA binding) and red (DNA binding or nucleotide binding). The bound nucleotide is colored pink. (c) Multiple sequence alignments with the same color code as defined above. Amino acid conservation with identical and similar (R/K, D/E, S/T, L/I/V/F/Y/W/M) residues is indicated by black and gray backgrounds, respectively. Secondary structure elements are schematically displayed over the sequences: helices and strands are shown by boxes and arrows, respectively. (d) Ribbon diagram representing superposition of the ATPase domains between RuvB (blue) and NSF (yellow), which have been classified into the AAA+ and AAA families, respectively.

Figure 2
Figure 2

Electron density maps and ribbon models of nucleotide-binding sites in the two ncs subunits. Possible residues that interact with nucleotides are depicted: Y14, I15, Y168, R179, and D180 are in contact with the adenine bases; K51 and T52 (Walker A), D97 (Walker B), T146 (Sensor I), and R205 (Sensor II) may interact with the phosphate groups. The stick models of (a) AMPPNP and (b) ADP were represented with corresponding simulated annealed Fo − Fc omit maps at a 1.5σ contour. The nucleotide atoms were omitted from the map calculation. Ribbons corresponding to the two sensor motifs and the two Walker motifs are indicated by the same color as in Fig. 1c. (c) Structural differences between the “A” (blue) and “B” (yellow) forms. Here, only the Cα backbones of domain N (ATPase domain) were superimposed between the two ncs molecules.

Figure 3
Figure 3

Characteristics of the RuvB molecular surface. (a) Electrostatic potential calculated with

grasp

(33) and (b) amino acid conservation, calculated with

clustalw

(34) and Toh's modified program (7) and based on 18 RuvB homologues, were mapped onto the

grasp

molecular surfaces by gradation coloring. Blue and red in a represent positively and negatively charged regions on a scale from −10.5 to +10.5, respectively, and white and red in b indicate low and high values of conservation, respectively. Perspectives are the same as in Fig. 1a on the Left and are from the rear on the Right.

Figure 4
Figure 4

Comparison of the hypothetical hexamer model of RuvB with the electron microscopic image. (a) Projection image (Left) of negative stained RuvB complexed with a 30-bp DNA, obtained by averaging 140 top views in our previous work (15). The resolution of the averaged image was 30.0 Å. The top views of the hexamer model (Center and Right) were constructed by superimposing each ATPase domain of RuvB (AMPPNP form) (blue region) onto the corresponding regions of HslU crystal structure (25) and the NSF crystal structure (23), respectively. The domains N, M, C, labeled residues, and the bound nucleotides are represented with the same color code as defined in Fig. 1. (b) Projection image (Left) of RuvB–DNA obtained by averaging 266 side views. This image of the single ring was taken from one-half of the double ring, which encircles duplex DNA. The resolution of the averaged image was 34.3 Å. Side view of the hexamer model (Right). [Reproduced with permission from ref. (Copyright 2000, Academic Press).]

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References

    1. Holliday R. Genet Res. 1964;5:282–304.
    1. Shinagawa H, Iwasaki H. Trends Biochem Sci. 1996;21:107–111. - PubMed
    1. West S C. Annu Rev Genet. 1997;31:213–244. - PubMed
    1. Tsaneva I R, Müller B, West S C. Cell. 1992;69:1171–1180. - PubMed
    1. Iwasaki H, Takahagi M, Nakata A, Shinagawa H. Genes Dev. 1992;6:2214–2220. - PubMed

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