Volume 12, Issue 10, Pages 1881-1889 (October 2004) A Novel Structure of DNA Repair Protein RecO from Deinococcus radiodurans  Nodar Makharashvili, Olga Koroleva, Sibes Bera, Duane P. Grandgenett, Sergey Korolev  Structure  Volume 12, Issue 10, Pages 1881-1889 (October 2004) DOI: 10.1016/j.str.2004.08.006

Figure 1 Ribbon Diagram of DrRecO and Surface Representation of the NTD and Its Interaction with the C Terminus of the Second Molecule (A) A ribbon representation of DrRecO structure. The protein is color-coded according to secondary structure elements, with numbered β strands (green) and lettered α helices (red). Zinc ion is shown as a cyan sphere, and zinc-coordinating cysteines are shown in stick representation. (B) Similar representation of DrRecO, but the molecule was rotated 90° around the vertical axis relative to the orientation in (A) (the N terminus is facing the viewer). (C) Surface representation of the NTD with the top part of the CTD in the same orientation as in (A) and color-coded according to the electrostatic potential, with blue color corresponding to positive charges and red to negative charges, calculated with program ICM. (D) Interaction of the C terminus of molecule A (shown as a stick representation in yellow) with the bottom of the β barrel of molecule B (shown as a molecular surface) observed in the crystal structure. Structure 2004 12, 1881-1889DOI: (10.1016/j.str.2004.08.006)

Figure 2 Multiple Alignment of 11 Distant RecO Sequences from Different Organisms Organisms are designated on the left of the alignment with the following abbreviations: Dr, D. radiodurans; Ec, E. coli; Bs, B. subtilis; Cp, C. pneumoniae; Fn, F. nucleatum; Mp, M. pulmonis; Rp, R. prowazekii; Sa, S. aureus; Tp, T. pallidum; Uu, Ureaplasma urealyticum; Xf, X. fastidiosa. Alignment was calculated with the program CLUSTLV (Pearson and Lipman, 1988), and the figure was prepared with the program ESPript2.0 (Gouet et al., 1999). Similar residues are highlighted in yellow, identical residues are in red, and zinc-coordinating cysteines of DrRecO and homologous cysteines in other sequences are highlighted by green. Secondary structure elements of DrRecO are shown above the sequences. Structure 2004 12, 1881-1889DOI: (10.1016/j.str.2004.08.006)

Figure 3 DNA-Annealing and Binding Properties of DrRecO and EcRecO (A) Annealing of complementary oligonucleotides. Two concentrations of both DrRecO and EcRecO proteins (10 nM and 100 nM) were used for measurements, and the concentration of DNA was 10 nM. The extent of DNA annealing was expressed as a percentage of the observed fluorescence quenching. (B) The ssDNA binding of DrRecO (dashed line) and EcRecO (solid line) to Fam-oligo1. The binding was monitored by measuring a change in fluorescence anisotropy of 10 nM solution of labeled oligonucleotide as a function of increasing protein concentrations. The results of titration were plotted as a relative anisotropy change. The initial phase of ssDNA binding (protein concentration less than 150 nM) is shown in the inset. (C) Similar measurements obtained with same ssDNA fragment annealed with complementary strand. Structure 2004 12, 1881-1889DOI: (10.1016/j.str.2004.08.006)

Figure 4 EcRecO Model (A) Ribbon and (B) electrostatic potential surface representation of the EcRecO structures in the same orientation as DrRecO in Figure 1A. (C) Electrostatic potential surface representation of DrRecO molecule A in a similar orientation. Structure 2004 12, 1881-1889DOI: (10.1016/j.str.2004.08.006)