1. The Human Cytomegalovirus UL44 C Clamp Wraps around DNA
Gloria Komazin-Meredith, Robert J. Petrella, Webster L. Santos, David J. Filman, James M. Hogle, Gregory L. Verdine, Martin Karplus, Donald M. Coen 
Structure 
Volume 16, Issue 8, Pages (August 2008)
DOI: /j.str
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2. Figure 1
Computational Model of the UL44 Processivity Factor Complexed with a DNA 12-Mer
The lowest-energy structure for the complex in the full-solvent (GBMV) calculation is shown in both a “C” view, in which the C-clamp shape of the protein is most readily seen (A), and a “back” view (B), which is generated by a 90° rotation of the C view about the vertical axis. The β strands of the protein are shown as cyan ribbons, the α helices are shown in red, and the DNA is shown as an orange double-helical backbone trace. The DNA molecule sits near the center of the cavity formed by the back faces of the monomers, with its longitudinal axis roughly (though not exactly) perpendicular to the plane of the dimer C. (The slight tilt of the DNA relative to the perpendicular is manifest in [B].) A connector loop, indicated in black, lies on the so-called front face of each UL44 monomer, joining its two halves; the back face of each monomer is opposite this face. The gap loops (residues 163–174), which are indicated in green in the upper monomer (monomer 1) and in magenta in the lower monomer (monomer 2), extend across the intermonomer gap and the periphery of the DNA (to its right in [A]). Lysine residues on the back face that were mutated in the biochemical studies (K35, K224, K158, and K237, from left to right in the upper monomer of [B]) are shown as black spheres. Cysteine residues targeted in the crosslinking studies are shown as yellow (C175) or magenta (C117) spheres.
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3. Figure 2 Protein-DNA Hydrogen Bonding and the Spiral Ionic Track
Stereoimages of the charged protein side chains of UL44 shown interacting with the DNA double-helix in the lowest-energy GBMV structure (A) and the second lowest-energy rdie structure (B). In this reduced representation of the DNA (indicated in orange), only the phosphate atoms of the DNA are shown explicitly, and the DNA has been shortened. The side chains from the cavity are shown in gray; those in the gap loops are shown in either green or magenta (consistent with Figure 1). The black dotted lines indicate hydrogen bonds with lengths of less than 3.5 Å (heavy atom to heavy atom) from any charged residue side-chain donor (in the binding cavity or on the flexible loops) to any DNA phosphate oxygen acceptor. The dark yellow lines indicate weaker hydrogen bonds (of between 3.5 and 5.5 Å in length) between charged side chains on the loops (not the cavity) and the DNA phosphate oxygens.
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4. Figure 3 Hydrogen-Bonding Pattern Involving the Gap Loops
Stereoimages of the loops at the intermonomer gap interacting with the DNA 12-mer for the lowest-energy GBMV structure (A) and the two lowest-energy rdie structures ([B], lowest; [C], second-lowest). The color schemes for the DNA and the side chains of the gap loops are the same as for the previous figures. All hydrogen bonds having a donor (of any atom type) in the loops at the intermonomer gap, acceptors in the phosphate oxygens, and lengths of <5.5 Å are indicated with dotted lines; those of <3.5 Å in length are indicated in black. The orientation of the views is rotated 180° in the plane of the page, relative to that in Figure 1B. The backbone atoms (oxygens excluded) of the loops are indicated in dark gray. For the DNA, only the phosphate atoms are shown explicitly.
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5. Figure 4 Gap Loop Conformations
In the modeled DNA/protein structures, the flexible loops on either side of the intermonomer gap tend to span the gap, but they do so in different conformations. The orientation of the views in A–E is the same as that in Figure 3. The backbones of the loops at the intermonomer gap are shown in gray (C) and blue (N), and the DNA is represented as in Figure 1.
(A and B) The two lowest-energy gap loop conformations from the GBMV calculations (loop backbones only) are shown.
(C and D) The gap loop conformations of the two lowest-energy rdie structures are shown.
(E and F) Two views of the five cluster centers (loop conformations that are closest to the calculated center of the clusters of low-energy structures) for the loop backbones in each monomer are shown; these panels give an indication of the general distribution of the low-energy structures. The view in (F), which demonstrates the “groove” formed by the structural distribution around the DNA backbone strand, is oriented obliquely and is related to the other views, in approximate terms, by a slight rotation of the complex about the vertical axis and an approximately 60° rotation about the longitudinal axis of the DNA.
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6. Figure 5 DNA Binding of Wild-Type and Mutant UL44 Proteins
The binding to DNA of (A) wild-type (WT) and basic, back face mutants (K35A, K158A, K224A, K237A, K35A/K158A/K224A/K237A), and (B) wild-type (WT) and gap loop mutants (R165A, K167A, R168A, K171A, K172A, R165A/K167A/R168A/K171A/K172A) was measured using a filter binding assay. Error bars represent standard errors of means from two to five experiments.
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7. Figure 6
Multiple-Alanine Substitution Mutants Retain Binding to the C-Terminal 22 Residues of UL54
(A) Raw isothermal titration calorimetry data for the titration of UL54 peptide into a sample cell containing wild-type or the indicated mutant UL44ΔC290 proteins.
(B) Amount of heat released per injection in kcal/mol of injectant plotted against the molar ratio of peptide to protein that is present in the cell at the time of each injection. The Kds calculated from the data are shown below the plots.
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8. Figure 7 UL44-DNA Crosslinking
(A) Chemical structure of the product of the crosslinking reaction (UL44 is in blue, N-thioalkyl phosphoramidite tether is in red, and DNA backbone is in green).
(B) Covalent complex formation of either wild-type or mutant UL44 proteins with thiol-tethered DNA analyzed on a nonreducing SDS-polyacrylamide gel. Also shown is a wild-type UL44 control with no DNA.
(C) DNA binding of wild-type and mutant UL44 proteins, measured by a filter binding assay. Error bars represent standard errors of means from two experiments.
Structure  , DOI: ( /j.str )
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