Binding of NHI and FX11 at the A-site.

<p>A) A representative MD snapshot of LDHA:NHI_A, with coloring scheme identical to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086365#pone-0086365-g006" target="_blank">Figure 6</a>. Black dashed lines represent polar interactions. B) Overlay of representative MD structures from four monomers of LDHA:FX11_A (carbon atoms in cyan, magenta, yellow, and pink, respectively) and PDB 1I10 (carbon atoms in grey). Grey surface indicates the solvent-accessible surface of LDHA, showing that two of the FX11 structures are completely outside the binding groove.</p

Structures of LDHA binders of interest.

<p>A-site and S-site binding moieties are indicated by boxes with blue dashed lines and red dashed lines, respectively.</p

Binding constants and site of binding of LDHA binders.

<p>^a The natural substrate pyruvate is referred to as PYR hereafter for simplicity.</p

Populations of important substrate binding interactions in simulations of monomeric vs tetrameric forms.

<p>^a Contacts are defined as any heavy atom pair with distance ≤0.4 nm.</p><p>^b O refers to the carbonyl oxygen of PYR, while OX2 refers to a carboxylate oxygen (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086365#pone.0086365.s005" target="_blank">Text S1</a>).</p

Binding of AJ1 and 1E7.

<p>Representative MD structures (cartoon and carbon atoms in cyan) and corresponding crystal structures (cartoon and carbon atoms in grey) are overlaid for A) LDHA:AJ1 and B) LDHA:1E7. Selected binding site residues are labeled and shown in thin lines, while ligands are shown in thick sticks. Dashed lines represent polar interactions. Other atoms are colored: oxygen, red; nitrogen, blue; sulfur, yellow; chlorine, green.</p

Structure of human LDHA (PDB 1I10).

<p>Amino acid residues are shown in cartoons and NADH/oxamate are shown in sticks. A) Tetrameric structure of human LDHA. Chains A, B, C, and D are colored green, yellow, magenta, and cyan, respectively. B) Close-up view of the binding site from chain A. The active site mobile loop is colored red.</p

Work and force involved in the pulling of LDHA binders from their binding sites.

<p>^a Calculated according to ΔG = −RTln(K_d) from experimental K_d values.</p><p>^b Reported as average ± standard deviation from 12 replicate steered MD runs.</p

Binding of NHI and FX11 at the S-site.

<p>Representative MD snapshots of A) LDHA:NHI_S and B) LDHA:FX11_S are shown. The color scheme is identical to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086365#pone-0086365-g006" target="_blank">Figure 6</a> with hydrogen atoms in white, while the mobile loop is in magenta.</p

Comparison of the binding of dual-site inhibitors.

<p>Representative MD snapshots of LDHA:0SN (carbon atoms in cyan) and LDHA:1E4 (carbon atoms in magenta) are superimposed. Selected binding site residues are labeled and shown in thin lines, while 0SN and 1E4 are shown in thick sticks. The mobile loop is represented by a ribbon, and the solvent-accessible surface of LDHA is indicated by a grey transparent surface. Dashed lines represent polar interactions. Other atoms are colored: oxygen, red; nitrogen, blue; sulfur, yellow; chlorine, green; fluorine, pale cyan.</p

Comparison of experimental binding free energies and calculated ΔPMF.

<p>^a Calculated according to ΔG = −RTln(K_d) from experimental K_d values.</p><p>^b Calculated by subtracting the ΔG_dissoc or ΔPMF of AJ1 for A-site binders.</p