Cumulative distributions of donor-acceptor distances determined for various types of intermolecular hydrogen bond donor-acceptor pairs identified in complexes of proteins with non-halogenated ligands, in which the ligand is either a hydrogen bond donor (A) or acceptor (B).

<p>Cumulative distributions of donor-acceptor distances determined for various types of intermolecular hydrogen bond donor-acceptor pairs identified in complexes of proteins with non-halogenated ligands, in which the ligand is either a hydrogen bond donor (A) or acceptor (B).</p

Comparison of distributions of hydrogen bond lengths, calculated separately for fluorinated (LF), otherwise halogenated (LX), and non-halogenated ligands (LH), for the four most represented topologies of protein-ligand hydrogen bonds.

<p>Those for which hydrogen bonds to LX/LF ligands are, according to the Mann-Whitney U test, significantly shorter (assuming α = 0.05) are highlighted. Note that for each pair of H-bond distributions, a smaller mean rank indicates statistically shorter donor-acceptor distances, or, equivalently, positive values of Z_U statistics indicate these types of H-bonds, which are shorter to halogenated ligands. The corresponding medians, and their differences with statistical significances (p), are also presented.</p

Cumulative distributions of NH•••O (blue) and O•••HN (red) intermolecular hydrogen bonds identified in protein complexes with non-halogenated (A, LH), fluorinated (B, LF) and otherwise halogenated ligands (C, LX).

<p>See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099984#pone-0099984-t002" target="_blank">Table 2</a> for details.</p

Results of the Kruskal-Wallis (K-W) test in the analysis of the topology-dependent length of a hydrogen bond between a non-halogenated ligand (LH) and a protein: for each pair of hydrogen bond acceptor/donor pair the p-value for the null hypothesis that both distributions are identical was estimated according to the two-tailed multiple comparison.

<p>The values marked in red denote the pairs of distributions that differ one from the other, with α = 0.05. Additionally, the identified number of each type of hydrogen bond, n, and mean rank test are presented.</p

Effect of a halogen atom on cumulative distributions determined for the four most abundant types of hydrogen bond donor-acceptor pairs: NH•••O (A), OH•••O (B), N•••HN (C), and O•••HN (D), respectively.

<p>The distributions estimated for non-halogenated (LH), fluorinated (LF) and other halogenated ligands (LX) are presented in black, blue and green, respectively. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0099984#pone-0099984-t003" target="_blank">Table 3</a> for details.</p

Occurrence of various types of hydrogen bonds identified in two groups of proteins (Enzyme Classification, EC, 2.3 or 2.7) for three types of ligands.

<p>These data include 41 hydrogen bonds to protein sulfur, which were excluded from further analyses.</p

Distribution of short halogen-acceptor (O, N, S, π system) contacts identified in 21 accessible structures of complexes of CK2α with halogenated ligands.

<p>The Gaussian cumulative distribution was fitted to the crystallographic data for halogen to donor distance (solid line in panel A) and, according to the Anderson-Darling test, experimental data up to 3.7 Å agrees with a normal distribution (panel B). However, pairs separated by more than 3.7 Å are overrepresented, clearly limiting the maximal distance for eventual halogen-bonding interactions to the sum of donor and acceptor VdW radii. Note that an isolated water molecule (red triangles in panels A, C) is an equally favorable acceptor to the protein (O, N, S, π-electrons). The cumulative distribution of the experimental data is visibly better represented by a bi-normal distribution (panel C), in which the contribution of an additional narrow peak represents putative halogen-bonding (panel D). This is additionally supported by the distribution of angles X…Acc-C and C-X…Acc, which, for short halogen-acceptor distances, are substantially restricted to the regions favoring halogen bond formation (see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0048898#pone.0048898.s002" target="_blank">Figure S2</a>).</p

Synthesis and Physico-Chemical Properties in Aqueous Medium of All Possible Isomeric Bromo Analogues of Benzo-1H-Triazole, Potential Inhibitors of Protein Kinases

In ongoing studies on the role of the individual bromine atoms of 4,5,6,7-tetrabromobenzotriazole (TBBt) in its relatively selective inhibition of protein kinase CK2α, we have prepared all the possible two mono-, four di-, and two tri-bromobenzotriazoles and determined their physicochemical properties in aqueous medium. They exhibited a general trend of a decrease in solubility with an increase in the number of bromines on the benzene ring, significantly modulated by the pattern of substitution. For a given number of attached bromines, this was directly related to the electronic effects resulting from different sites of substitution, leading to marked variations of p<i>K</i>_a values for dissociation of the triazole proton. Experimental data (p<i>K</i>_a, solubility) and <i>ab initio</i> calculations demonstrated that hydration of halogenated benzotriazoles is driven by a subtle balance of hydrophobic and polar interactions. The combination of QM-derived free energies for solvation and proton dissociations was found to be a reasonably good predictor of inhibitory activity of halogenated benzotriazoles vs CK2α. Since the pattern of halogenation of the benzene ring of benzotriazole has also been shown to be one of the determinants of inhibitory potency vs some viruses and viral enzymes, the present comprehensive description of their physicochemical properties should prove helpful in efforts to elucidate reaction mechanisms, including possible halogen bonding, and the search for more selective and potent inhibitors

Schematic representation of the effects of stepwise bromination of the benzene ring of benzotriazole (Bt), including interdependence of the molecular volume (A) and pK_a (B) of the products with their IC₅₀ for inhibition of protein kinase CK2α: (a) Green arrows follow sequential bromination of the central vicinal C(5)/C(6) atoms, leading to a moderate decrease of pK_a and a large decrease in IC₅₀; (b) Red arrows illustrate the lesser effects of bromination of the peripheral C(4)/C(7) atoms, resulting in a significant decrease of pK_a, with virtually no gain in inhibitory activity; (c) Black lines link products with the same number of bromine atoms, but substantially differing in pK_a and IC₅₀, culminating in the di- and tri-brominated derivatives with inhibitory activities comparable to that of TBBt.

<p>Note that the parent Bt (pKa 8.56), which is not an inhibitor (IC₅₀>2 mM), is located outside the diagrams.</p

Structures of all possible halogenated derivatives of benzotriazole.

<p>Structures of all possible halogenated derivatives of benzotriazole.</p