Use of biopartitioning micellar chromatography and RP-HPLC for the determination of blood-brain barrier penetration of α-adrenergic/imidazoline receptor ligands, and QSPR analysis.

For this study, 31 compounds, including 16 imidazoline/α-adrenergic receptor (IRs/α-ARs) ligands and 15 central nervous system (CNS) drugs, were characterized in terms of the retention factors (k) obtained using biopartitioning micellar and classical reversed phase chromatography (log kBMC and log kwRP, respectively). Based on the retention factor (log kwRP) and slope of the linear curve (S) the isocratic parameter (φ0) was calculated. Obtained retention factors were correlated with experimental log BB values for the group of examined compounds. High correlations were obtained between logarithm of biopartitioning micellar chromatography (BMC) retention factor and effective permeability (r(log kBMC/log BB): 0.77), while for RP-HPLC system the correlations were lower (r(log kwRP/log BB): 0.58; r(S/log BB): -0.50; r(φ0/Pe): 0.61). Based on the log kBMC retention data and calculated molecular parameters of the examined compounds, quantitative structure-permeability relationship (QSPR) models were developed using partial least squares, stepwise multiple linear regression, support vector machine and artificial neural network methodologies. A high degree of structural diversity of the analysed IRs/α-ARs ligands and CNS drugs provides wide applicability domain of the QSPR models for estimation of blood-brain barrier penetration of the related compounds.

Polypharmacology of dopamine receptor ligands.

Most neurological diseases have a multifactorial nature and the number of molecular mechanisms discovered as underpinning these diseases is continuously evolving. The old concept of developing selective agents for a single target does not fit with the medical need of most neurological diseases. The development of designed multiple ligands holds great promises and appears as the next step in drug development for the treatment of these multifactorial diseases. Dopamine and its five receptor subtypes are intimately involved in numerous neurological disorders. Dopamine receptor ligands display a high degree of cross interactions with many other targets including G-protein coupled receptors, transporters, enzymes and ion channels. For brain disorders like Parkinsons disease, schizophrenia and depression the dopaminergic system, being intertwined with many other signaling systems, plays a key role in pathogenesis and therapy. The concept of designed multiple ligands and polypharmacology, which perfectly meets the therapeutic needs for these brain disorders, is herein discussed as a general ligand-based concept while focusing on dopaminergic agents and receptor subtypes in particular.

C-terminal truncation of yeast SerRS is toxic for Saccharomyces cerevisiae due to altered mechanism of substrate recognition.

Like all other eukaryal cytosolic seryl-tRNA synthetase (SerRS) enzymes, Saccharomyces cerevisiae SerRS contains a C-terminal extension not found in the enzymes of eubacterial and archaeal origin. Overexpression of C-terminally truncated SerRS lacking the 20-amino acid appended domain (SerRSC20) is toxic to S. cerevisiae possibly because of altered substrate recognition. Compared to wild-type SerRS the truncated enzyme displays impaired tRNA-dependent serine recognition and is less stable. This suggests that the C-terminal peptide is important for the formation or maintenance of the enzyme structure optimal for substrate binding and catalysis.

Dynamic effects of food consistency on chewing motions.

The purpose of the study was to find evidence of how different types of food consistency affect chewing motions, especially the forward, downward and sidewise extents of motion of the lower jaw. Nineteen individuals with intact tooth sequence, aged from 20 to 37 years, were asked to chew three types of food of different consistency (banana, bread, carrot). The motions of the lower jaw were recorded by ELITE system, i.e. the measurement instrument that by stereo-photo-grametric procedures calculates space co-ordinates of markers on faces of the study subjects. The system enables continuous recording of lower jaw motions in three dimensions, without any possibility of the study subjects' influencing the operation of the instrument, which significantly decreases the possibility of error. Study results have shown that in all 19 subjects a greater food consistency increases the extent of chewing motion. In each individual study subject different average values were found for equal shifts of lower jaw when chewing the same type of food. Although varying from subject to subject, the chewing cycle depends to a great extent on food consistency. By increasing the consistency of a bite, the extent of lower jaw motion has increased in every single study subject.

QSAR study of selective I1-imidazoline receptor ligands.

Selective imidazoline(1)-receptor (I(1)-R) ligands are used clinically to reduce blood pressure. Thus, there is significant interest in developing new imidazoline analogs with high selectivity and affinity for I(1) receptors. A quantitative structure-activity relationship (QSAR) study was carried out on 11 potent I(1)-R ligands (derivatives of imidazoline, oxazoline and pyrroline) using a multiple linear regression (MLR) procedure. The selected compounds have been studied using B3LYP/3-21G(d, p) and B3LYP/6-31G(d, p) methods. Among the 42 descriptors that were considered in generating the QSAR model, three descriptors (partial atomic charges of nitrogen in the heterocyclic moiety (N-2 charge), log D and the dipole moment of the ligands) resulted in a statistically significant model with r(2) > 0.874 and [image omitted] > 0.802. The QSAR models were validated through cross-validation and external test set prediction. The aim of the developed MLR models for the I(1)-R ligands was to link the structures to their reported I(1)-R binding affinity log(1/Ki). The proposed QSAR models indicate that an increase in log D and the dipole moment value and a decrease in N-2 charge in the heterocyclic moiety are predictors of better selectivity and affinity for I(1) receptors. The developed QSAR model is intended to predict the I(1)-R binding affinity of related compounds and aid in the rational design of new potent and selective I(1)-R ligands.

Dynamic influence of food consistency on the masticatory motion.

The masticatory cycle is a complex process and it depends on many factors. In our study we wanted to prove to what extent various types of food consistency influence the masticatory motions, especially the extent of forward, downward and lateral motions of the mandible. Nineteen study subjects aged from 20 to 37 years and with intact teeth rows were asked to chew three types of food of various consistency (banana, bread and carrot). The motions of the mandible were recorded by stereo-photo-grammetric system. Study results have shown that in all 19 study subjects the increase in food consistency increases the extent of masticatory motions. The average size of forward mandibular motion in all 19 study subjects when chewing banana amounts to 2.65 mm, when chewing bread it is 2.96 mm and 3.64 mm when chewing carrot. The average size of downward mandibular motion for all 19 study subjects amounts to 6.79 mm when chewing banana, 7.17 mm when chewing bread and 8.09 mm when chewing carrot. The average size of lateral mandibular motion in all 19 study subjects amounts to 2.46 mm when chewing banana, 2.80 mm when chewing bread and 3.40 mm when chewing carrot. Although varying from subject to subject, the masticatory cycle significantly depends on food consistency. By increasing the consistency of a mouthful, the extent of mandibular motion increases in every single study subject.

The C-terminal extension of yeast seryl-tRNA synthetase affects stability of the enzyme and its substrate affinity.

Saccharomyces cerevisiae seryl-tRNA synthetase (SerRS) contains a 20-amino acid C-terminal extension, which is not found in prokaryotic SerRS enzymes. A truncated yeast SES1 gene, lacking the 60 base pairs that encode this C-terminal domain, is able to complement a yeast SES1 null allele strain; thus, the C-terminal extension in SerRS is dispensable for the viability of the cell. However, the removal of the C-terminal peptide affects both stability of the enzyme and its affinity for the substrates. The truncation mutant binds tRNA with 3.6-fold higher affinity, while the Km for serine is 4-fold increased relative to the wild-type SerRS. This indicates the importance of the C-terminal extension in maintaining the overall structure of SerRS.

Defining the active site of yeast seryl-tRNA synthetase. Mutations in motif 2 loop residues affect tRNA-dependent amino acid recognition.

The active site of class II aminoacyl-tRNA synthetases contains the motif 2 loop, which is involved in binding of ATP, amino acid, and the acceptor end of tRNA. In order to characterize the active site of Saccharomyces cerevisiae seryl-tRNA synthetase (SerRS), we performed in vitro mutagenesis of the portion of the SES1 gene encoding the motif 2 loop. Substitutions of amino acids conserved in the motif 2 loop of seryl-tRNA synthetases from other sources led to loss of complementation of a yeast SES1 null allele strain by the mutant yeast SES1 genes. Steady-state kinetic analyses of the purified mutant SerRS proteins revealed elevated Km values for serine and ATP, accompanied by decreases in kcat (as expected for replacement of residues involved in aminoacyl-adenylate formation). The differences in the affinities for serine and ATP, in the absence and presence of tRNA are consistent with the proposed conformational changes induced by positioning the 3'-end of tRNA into the active site, as observed recently in structural studies of Thermus thermophilus SerRS (Cusack, S., Yaremchuk, A., and Tukalo, M. (1996) EMBO J. 15, 2834-2842). The crystal structure of this moderately homologous prokaryotic counterpart of the yeast enzyme allowed us to produce a model of the yeast SerRS structure and to place the mutations in a structural context. In conjunction with structural data for T. thermophilus SerRS, the kinetic data presented here suggest that yeast seryl-tRNA synthetase displays tRNA-dependent amino acid recognition.