Molecular docking study of the binding of aminopyridines within the K+ channel.

We present a molecular docking study aimed to identify the binding site of protonated aminopyridines for the blocking of voltage dependent K(+) channels. Several active aminopyridines are considered: 2-aminopyridine, 3-aminopyridine, 4-aminopyridine, 3,4-diaminopyridine, and 4-aminoquinoleine. We apply the AutoDock force field with a lamarckian genetic algorithm, using atomic charges for the ligands derived from the electrostatic potential obtained at the B3LYP/cc-pVDZ level. We find a zone in the alpha-subunit of the K(+) channel bearing common binding sites. This zone corresponds to five amino acids comprised between residuals Thr107 and Ala111, in the KcsA K(+) channel (1J95 pdb structure). The 2-aminopyridine, 3-aminopyridine, 4-aminopyridine, and 3,4-diaminopyridine bind to the carboxylic oxygens of Thr107 and Ala111. In all cases aminopyridines are perpendicular to the axis of the pore. 4-aminoquinoleine binds to the carboxylic oxygen of Ala111. Due to its large size, the molecular plane is parallel to the axis of the pore. The charge distributions and the structures of the binding complexes suggest that the interaction is driven by formation of several hydrogen bonds. We find 2-aminopyridine, 3-aminopyridine, 4-aminopyridine, and 3,4-diaminopyridine with similar binding energy. Considering the standard error of the estimate of the AutoDock force field, this energy should lie, as a rough estimation, in the interval 3-7 kcal mol(-1). On the other hand, 4-aminoquinoleine seems to have a smaller binding energy.

Conformations, protonation sites, and metal complexation of benzohydroxamic Acid. A theoretical and experimental study.

A theoretical and experimental study on the structure and deprotonation of benzohydroxamic acid (BHA) has been performed. Calculations at the RHF/cc-pVDZ level, refined by the B3LYP/AUG-cc-pVDZ method, indicate that, in the gas phase, Z amide is the most stable structure of both neutral and deprotonated BHA. (1)H-(1)H nuclear Overhauser enhancement spectroscopy and (1)H-(1)H correlation spectroscopy spectra in acetone, interpreted with ab initio interatomic distances, reveal that BHA is split into the Z and E forms, the [E]/[Z] ratio being 75:25 at -80 degrees C. The formation of E-E, Z-Z, and E-Z dimers has been detected; in the presence of water, the dimers dissociate to the corresponding monomers. The rates of proton exchange within the Z and E forms and between E and Z were measured by dynamic (1)H NMR in the -60 to 40 degrees C temperature range; an increase in water content lowers the rate of exchange of the E isomer. The effect of D(2)O on the NMR signals indicates a fast hydrogen exchange between D(2)O and the E and Z amide forms. The sequence of the acid strength at low temperatures is (N)H(E)) approximately (O)H(E) < (O)H(Z) approximately (N)H(Z). The kinetics of complex formation between BHA and Ni(2+), investigated by the stopped-flow method, show that both neutral BHA and its anion can bind Ni(2+). Whereas the anion reacts at a "normal" speed, the rate of water replacement from Ni(H(2)O)(6)(2+) by neutral BHA is about 1 order of magnitude less than expected. This behavior was interpreted assuming that, in aqueous solution, BHA mainly adopts a closed (hydrogen-bonded) Z configuration, which should open (with an energy penalty) for the metal binding process to occur.

Theoretical analysis of pyridine protonation in water clusters of increasing size.

The protonation of pyridine in water clusters as a function of the number of water molecules was theoretically analyzed as a prototypical case for the protonation of organic bases. We determined the variation of structural, bonding, and energetic properties on protonation, as well as the stabilization of the ionic species formed. Thus, we used supermolecular models in which pyridine interacts with clusters of up to five water molecules. For each complex, we determined the most stable unprotonated and protonated structures from a simulated annealing at the semi ab initio level. The structures were optimized at the B3LYP/cc-pVDZ level. We found that the hydroxyl group formed on protonation of pyridine abstracts a proton from the ortho-carbon atom of the pyridine ring. The "atoms in molecules" theory showed that this C-H group loses its covalent character. However, starting with clusters of four water molecules, the C-H bond recovers its covalent nature. This effect is associated with the presence of more than one ring between the water molecules and pyridine. These rings stabilize, by delocalization, the negative charge on the hydroxyl oxygen atom. Considering the protonation energy, we find that the protonated forms are increasingly stabilized with increasing size of the water cluster. When zero-point energy is included, the variation follows closely an exponential decrease with increasing number of water molecules. Analysis of the vibrational modes for the strongest bands in the IR spectra of the complexes suggests that the protonation of pyridine occurs by concerted proton transfers among the different water rings in the structure. Symmetric water stretching was found to be responsible for hydrogen transfer from the water molecule to the pyridine nitrogen atom.

Mechanism of pyridine protonation in water clusters of increasing size.

This work presents a theoretical mechanistic study of the protonation of pyridine in water clusters, at the B3LYP/cc-pVDZ theory level. Clusters from one to five water molecules were used. Starting from previously determined structures, the reaction paths for the protonation process were identified. For complexes of pyridine with water clusters of up to three water molecules just one transition state (TS) links the solvated and protonated forms. It is found that the activation energy decreases with the number of water molecules. For complexes of four and five water molecules two transition states are found. For four water molecules, the first TS links the starting solvated structure with a new, less stable, solvated form through a concerted proton transfer between a ring of water molecules. The second TS links the new solvated structure to the protonated form. Thus, protonation is a two-step process. For the five water molecules cluster, the new solvated structure is more stable than the starting one. This structure exhibits two double hydrogen bonds involving the pyridinic nitrogen and several water molecules. The second TS links the new structure with the protonated form. Now the process occurs in one step. In all cases considered, the proton transfers involve an interconversion between covalent and hydrogen bonds. For four and five water molecules, the second TS is structurally and energetically very close to the protonated form. As evidenced by the vibration frequencies, this is due to a flat potential energy hypersurface in the direction of the reaction coordinate. Determination of DeltaG at 298.15 K and 1 atm shows that the protonation of pyridine needs at least four water molecules to be spontaneous. The complex with five water molecules exhibits a large DeltaG. This value yields a pKa of 2.35, relatively close to the reported 5.21 for pyridine in water.

Deprotonation sites of acetohydroxamic acid isomers. A theoretical and experimental study.

Theoretical (ab initio calculations) and experimental (NMR, spectrophotometric, and potentiometric measurements) investigations of the isomers of acetohydroxamic acid (AHA) and their deprotonation processes have been performed. Calculations with the Gaussian 98 package, refined at the MP2(FC)/AUG-cc-pVDZ level considering the molecule isolated, indicate that the Z(cis) amide is the most stable form of the neutral molecule. This species and the less stable (Z)-imide form undergo deprotonation, giving rise to two stable anions. Upon deprotonation, the E(trans) forms give three stable anions. The ab initio calculations were performed in solution as well, regarding water as a continuous dielectric; on the basis of the relative energies of the most stable anion and neutral forms, calculated with MP2/PCM/AUG-cc-pVDZ, N-deprotonation of the amide (Z or E) structure appeared to be the most likely process in solution. NMR measurements provided evidence for the existence of (Z)- and (E)-isomers of both the neutral and anion forms in solution. Comparisons of the dynamic NMR and NOESY (one-dimensional) results obtained for the neutral species and their anions were consistent with N-deprotonation, which occurred preferentially to O-deprotonation. The (microscopic) acid dissociation constants of the two isomers determined at 25 degrees C from the pH dependence of the relevant chemical shifts, pK(E) = 9.01 and pK(Z) = 9.35, were consistent with the spectrophotometric and potentiometric evaluations (pK(HA) = 9.31).

Rational modelling of the voltage-dependent K+ channel inactivation by aminopyridines.

A functional model for the in vitro inactivation of voltage-dependent K(+) channels is developed. The model expresses the activity as a function of the aminopyridine pK(a), the interaction energy with the receptor, and a quotient of partition functions. Molecular quantum similarity theory is introduced in the model to express the activity as a function of the principal components of the similarity matrix for a series of agonists. To validate the model, a set of five active (protonated) aminopyridines is considered: 2-aminopyridine, 3-aminopyridine, 4-aminoquinoleine, 4-aminopyridine, and 3,4-diaminopyridine. A regression analysis of the model gives good results for the variation of the observed activity with the overlap similarity index when pyridinic rings are superposed. The results support the validity of the model, and the hypothesis of a ligand-receptor entropy variation depending mainly on the nature of the ligand. In addition, the results suggest that the pyridinic ring must play an active role in the interaction with the receptor site. This interaction with the protonated pyridinic nitrogen can involve a cation-pi interaction or a donor hydrogen bond. The amine groups, at different relative positions of the pyridinic nitrogen, can form one or more hydrogen bonds due to the C(4) symmetry of the inner part of the pore in the K(+) channel.

The nature of the receptor site for the reversible K+ channel blocking by aminopyridines.

This work presents a theoretical study aimed at the identification of the receptor site for the blocking of the voltage dependent K+ channels by protonated aminopyridines. Thus, the density functional theory (DFT) at the B3LYP/6-311 G (d,p) level is applied, both in vacuum and in solution, to a series of active (protonated) compounds: 2-aminopyridine, 3-aminopyridine, 4-aminopyridine, 3,4-diaminopyridine, and 4-aminoquinoleine. Analysis of the X-ray structure of the alpha-subunit of the channel shows that charged aminopyridines can interact electrostatically with a glutamic acid residue in the outside of the pore, or through a cation-pi interaction in the inside. To test both possibilities, model complexes are built using as nucleophiles a carboxylic group and an ethylene molecule, respectively. The three-dimensional electrostatic potential distribution of the protonated aminopyridines shows that an approaching nucleophile will be oriented toward the N-H (protonated) bond. Interaction with the carboxylic residue leads to a proton transfer, with the aminopyridine-carboxylic acid linked by a hydrogen bond. The observed breaking of the equivalence of the Laplacian of the charge density, the relative energy variation for the complexes, and the interaction with only one of the carboxylic residues in the fourfold alpha-subunit of the K+ channel are not compatible with the observed in vitro activity variation of aminopyridines. On the other hand, the study on the ethylene complexes shows, in vacuum and solution, a cation-pi interaction, clearly characterized by the atoms in molecules (AIM) theory. The variation of relative energy in solution is very small, but approaches the variation of in vitro activity. Our results, the pharmacophoric characteristics of aminopyridines, and the analysis of the three-dimensional internal structure of the K+ channel alpha-subunit suggest two putative receptor sites. One is formed by the four Thr-Thr-Val chains conforming the entrance to the narrow part of the inner K+ channel. The other is defined by four Thr residues within the pore.