1. Volume 20, Issue 1, Pages 155-166 (October 2005)
MutH Complexed with Hemi- and Unmethylated DNAs: Coupling Base Recognition and DNA Cleavage 
Jae Young Lee, Judy Chang, Nimesh Joseph, Rodolfo Ghirlando, Desirazu N. Rao, Wei Yang 
Molecular Cell 
Volume 20, Issue 1, Pages (October 2005)
DOI: /j.molcel
Copyright © 2005 Elsevier Inc. Terms and Conditions

2. Figure 1 Ribbon Diagrams of the MutH-DNA Complexes
(A) Orthogonal views of the unmethylated complex. The active site residues are shown as pink ball-and-stick diagrams, and the residues involved in sequence-specific DNA recognitions are shown as blue ball-and-stick diagrams. The Ca2+ ions are shown as green spheres.
(B) The hemimethylated complex. The GATC sequences are highlighted in orange and the methylated Ade is shown in red.
Molecular Cell  , DOI: ( /j.molcel )
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3. Figure 2 Structural Comparisons of MutH with and without DNA
(A) Stereo view of superimposed Cα traces of three apo structures and three DNA complexes. The apo structures are shown in light (1AZO), medium (2AZO, chain A), and dark yellow-orange (2AZO, chain B), and MutHs complexed with DNA are shown in blue (unmethylated) and purple (hemimethylated).
(B) Superposition of the six Cα traces of the N-arm subdomain.
(C) Topology diagram of MutH. The catalytic core and the DNA binding module are highlighted in pink and blue, respectively, and the active site and other critical residues are labeled.
(D) Side-by-side ribbon diagrams of MutH and MutL (LN40). The interaction regions identified by protein crosslinking are shaded in blue ovals.
Molecular Cell  , DOI: ( /j.molcel )
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4. Figure 3 The DNAs and DNA-MutH Interactions
(A) Superposition of the hemi- (orange) and unmethylated (yellow) DNA complexed with MutH. The backbone of an ideal B-form DNA (semi-transparent gray) is included for reference. The methyl-adenine is highlighted in red. The Cα trace of the region responsible for communicating methyl detection to the active site (highlighted in Figure 2C) is shown, and the C1 loop is highlighted in green (unmethylated) and pink (hemimethylated). The side chains of F94, Y212, the DEK motif, and S65 are shown in ball-and-stick models. Nitrogen and oxygen atoms are shown as blue and red spheres, respectively.
(B) A diagram of interactions between MutH and hemi and unmethylated DNA. Blue labels indicate interactions by protein main chains, red by side chains, and green by both. Yellow highlight indicates interactions observed in both complexes, green in unmethylated complexes, and pink in the hemimethylated complex. Major-groove interactions are placed above each base, and minor-groove interactions are placed below. Solid lines indicate hydrogen bonds, and dashed lines indicate van der Waals contacts. Water-mediated interactions are labeled with “W.”
(C) Conformational changes between the hemi (pink) and unmethylated (green) complexes around the C1 loop.
(D) A close up of the molecular surface of MutH that complements the methylated Ade. R184, P185, and Y212 and the corresponding surface are highlighted in blue.
(E) Sequence-specific interactions between MutH and hemimethylated GATC in the major groove. A 2Fo-Fc map is superimposed on the DNA. The dotted lines in C, D, and E represent hydrogen bonds.
Molecular Cell  , DOI: ( /j.molcel )
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5. Figure 4 DNA Binding and Cleavage by MutH
(A) Sedimentation equilibrium profiles of unmethylated (green), hemi (pink), and fully methylated (blue) DNA with H. infl MutH at 1:1 molar ratio (3 μM each) plotted as a distribution of ln(A280) versus r2 at equilibrium. The dark blue straight line represents the expected curve for the noninteracting 1:1 mixture of the DNA and MutH, and the red line represents the perfect 1:1 complex. The residuals obtained from an unweighted single ideal solute best-fit analysis are shown on the right.
(B) Comparison of the active site configuration in the unmethylated (left) and the hemimethylated complex (right). Metal-ion coordination is represented by green dotted lines, and hydrogen bonds are depicted by red dotted lines. The double arrowhead and distance measurements indicate the largest changes between the two complexes.
(C) Time course of DNA cleavage by MutH. Hemi or unmethylated 12 bp DNA (2.0 µM) was incubated with MutH (0.1 µM) for the indicated time period, and cleavage products were resolved by electrophoresis on a 20% TBE urea gel. DNA cleavage in three measurements is averaged and plotted on the right. Error bars represent deviations among the three measurements.
(D) Cleavage of hemimethylated GATC with the Sp or Rp phosphorothioate substitution 3′ to the scissile bond. The DNA is 2.0 µM and MutH is 0.02 µM. Increasing MutH concentration does not improve cleavage of the Sp substrate.
Molecular Cell  , DOI: ( /j.molcel )
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6. Figure 5 The Active Site of MutH, BglI, BamHI, and BglII
DNA is shown as yellow sticks, with the scissile phosphate and its 3′ neighbor highlighted. The protein Cα traces surrounding each active site are shown as pink tubes. The metal ions (green), the active site residues, the carbonyl between the E and K of the DEK motif, and the nucleophilic water (red) are shown in ball-and-stick models. For clarity, other water molecules coordinating the metal ions are removed after the metal ion coordination is determined and are shown as green dashed lines. Hydrogen bonds are shown as red dotted lines. The pro-Sp and pro-Rp oxygen are labeled as Sp and Rp, respectively.
Molecular Cell  , DOI: ( /j.molcel )
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7. Figure 6 The Linchpin Model
The third residue in the catalytic triad, Lys, Glu, or Gln (highlighted in red), links the active site (indicated by green metal ions and pink carboxylates) and the DNA binding module (blue) via a hydrogen bond network (dotted red lines). The Cα trace of the catalytic core is shown in pink, the DNA binding module is shown in blue, and the linker in between is depicted in purple. DNAs are shown as space-filling models with backbones highlighted. The Cα traces of the neighboring subunit in the BamHI and BglII dimer are shown as lilac tubes. The side chains that made sequence-specific contact with DNA are shown as cyan ball-and-stick models with red oxygen and blue nitrogen atoms.
Molecular Cell  , DOI: ( /j.molcel )
Copyright © 2005 Elsevier Inc. Terms and Conditions