Identification of potential drug targets and inhibitor of the pathogenic bacteria Shigella flexneri 2a through the subtractive genomic approach.

Shigella flexneri 2a is one of the most pathogenic bacteria among the Shigella spp., which is responsible for dysentery and causes masses of deaths throughout the world per year. A proper identification of the potential drug targets and inhibitors is crucial for the treatment of the shigellosis due to their emerging multidrug resistance (MDR) patterns. In this study, a systematic subtractive approach was implemented for the identification of novel therapeutic targets of S. flexneri 2a (301) through genome-wide metabolic pathway analysis of the essential genes and proteins. Ligand-based virtual screening and ADMET analyses were also made for the identification of potential inhibitors as well. Initially, we found 70 essential unique proteins as novel targets. After subsequent prioritization, finally we got six unique targets as the potential therapeutic targets and their three-dimensional models were built thereafter. Aspartate-ß-semialdehyde dehydrogenase (ASD), was the most potent target among them and used for docking analysis through ligand-based virtual screening. The compound 3 (PubChem CID: 11319750) suited well as the best inhibitor of the ASD through ADMET and enzyme inhibition capacity analysis. To end, we hope that our proposed therapeutic targets and its inhibitors might give some breakthrough to treat shigellosis efficiently in in vitro.

In vitro screening for inhibitor of cloned Drosophila melanogaster tyramine-ß-hydroxylase and docking studies.

Biogenic amines are common biologically active substances extended within the whole animal kingdom where they play vital roles as signal transducer as well as regulator of cell functions. One of these biogenic amines called octopamine (OA) is synthesized from tyramine (TA) by the catalysis of tyramine-ß-hydroxylase (TßH) originated in the insect nervous system. Both TA and OA act as neurotransmitters, neurohormones and neuromodulators in the arthropod nervous system. Herein, the inhibitory activity of 1-arylimidazole-2(3H)-thiones (AITs) was tested on cloned Drosophila tyramine-ß-hydroxylase (DmTßH) expressed in Bombyx mori strain. Radiolabelled 3H-TA was used to analyze the activity of AITs exhibited inhibitory effects on DmTßH, whose ID50 values range from 0.02 to 2511nM where DmTßH was inhibited in a dose-dependent manner at pH 7.6 and 25°C during a 30min of incubation. To understand the catalytic role of the TßH, a three dimensional structure of the TßH from Drosophila melanogaster was constructed by homology modeling using the Phyre2 web server with 100% confidence. The modeled three-dimensional structure of TßH was used to perform the docking study with AITs. This may give more insights to precise design of inhibitors for TßH to control insect's population.

A randomized ethnomedicinal survey of snakebite treatment in southwestern parts of Bangladesh.

Snakebite is the single most important toxin-related injury, causing substantial mortality in many parts of the Africa, Asia and the Americas. Incidence of snakebite is usually recorded in young people engaged in active physical work in rural areas. The various plant parts used to treat snakebite included whole plant, leaves, barks, roots and seeds. Most bites in Bangladesh are recorded between May and October with highest number in June. Lower and upper limbs are most common sites of snakebite, but it may happen in other sites as well. Snake venom ( shé dú) has been the cause of innumerable deaths worldwide. However, antiserum does not provide enough protection against venom induced hemorrhage, necrosis, nephrotoxicity and hypersensitivity reactions. Informed consent was obtained from the practitioners prior to interviews. After the survey, it is concluded that the medicinal plants used by tribal medicinal practitioners in Bangladesh for treatment against snakebite are Acyranthes aspera L. ( tǔ niú xi), Amaranthus Viridis L. ( ye xiàng cài), Asparagus racemosus Willd ( zǒng xù tian dong) and Emblica officinalis Gaertn ( yóu gan), while the non-tribal communities used 35 plant species among them, most of the plants reported as new species used against snakebite in the belonging family. The plants present a considerable potential for discovery of novel compounds with fewer side effects for treatment of antisnake venom and can, at least in Bangladesh, become a source of affordable and more easily available drugs.

Homology modeling, agonist binding site identification, and docking in octopamine receptor of Periplaneta americana.

AY333178 (from Periplaneta americana, 628 AAs) was selected as a target octopamine receptor (OAR) class OAR2 for this study using Discovery Studio (DS Modeling1.1/1.2, Accelrys Inc.). Blast similarity search was performed and identified that AY333178 contains N-terminal domain of GPCR. Based upon Blast and Pfam results, Rhodopsin 1U19 (protein data bank) was considered as an ideal homologue and used as a template for homology modeling due to its higher X-ray resolution at 2.2A. Sequence alignment between AY333178 and 1U19 was done using Align123 followed by a manual modification. The final alignment was carefully evaluated and evidenced to be matching the conserved residue data for class A GPCR fairly well. The 3D model of AY333178 was generated with MODELER, and further refined using CHARMm. Superimposition of the model was done over the template 1U19. Two fairly consistent profiles were observed demonstrating AY333178 model was reasonable and could be employed for the further docking study. Agonist docking into OAR2 model was done using LigandFit. The superimposition of two top poses of representative agonists was performed with a soft surface generated. Those models are considered to be used in designing new leads for hopefully more active compounds. Further research on the comparison of models for the agonists may elucidate the mechanisms of OAR2-ligand interactions.

Effect of tyramine and stress on sex-pheromone production in the pre- and post-mating silkworm moth, Bombyx mori.

Tyramine (TA) increased significantly after mating, whereas there were no significant differences in octopamine (OA) and dopamine (DA) levels in the brain-suboesophageal ganglion (SOG) complexes between virgin and mated females. The effects of various biogenic amines were tested on pheromone production of virgin and mated females of the silkworm moth, Bombyx mori. After 8h a significant reduction by TA (46%) was observed. Meanwhile, when OA or DA was injected, a significant increase of pheromone titer was observed in both virgin and mated females. This study also presents evidence for an increase in levels of OA and DA in the brain-SOG complexes in response to mechanical stress in B. mori female. TA suppressed pheromone production in an in vitro pheromone gland (PG) homogenate preparation, thus suggesting that the target of TA is the PG. TA inhibited pheromone production in vitro in a dose-dependent manner and DA had a lower inhibitory activity than TA, whereas OA had no effect, suggesting that TA is a candidate for regulating pheromone production in the PG, although other factors could be responsible for the pheromonostatic function.

The pheromone production of female Plodia interpunctella is inhibited by tyraminergic antagonists.

Several compounds were found to suppress the calling behavior and in vitro pheromone biosynthesis of the Indian meal moth, Plodia interpunctella. The compounds were screened by means of a calling-behavior bioassay with female P. interpunctella. Five derivatives with activities in the nanomolar range were identified, in order of decreasing pheromonostatic activity: 4-hydroxybenzaldehyde semicarbazone (42) > 5-(4-methoxyphenyl)-1,3-oxazole (38) > 5-[4-(tert-butyl)phenyl]-1,3-oxazole (40) > 5-(3-methoxyphenyl)-1,3-oxazole (35) > 5-(4-cyanophenyl)-1,3-oxazole (36). These compounds also showed in vitro inhibitory activity in intracellular de novo pheromone biosynthesis, as determined with isolated pheromone-gland preparations that incorporated [1-(14)C]sodium acetate in the presence of the so-called pheromone-biosynthesis-activating neuropeptide (PBAN). The non-additive effect of the inhibitor with antagonist (yohimbine) for the tyramine (TA) receptor suggests that it could be a tyraminergic antagonist. Three-dimensional (3D) computer models were built from a set of compounds. Among the common-featured models generated by the program Catalyst/HipHop, aromatic-ring (AR) and H-bond-acceptor-lipophilic (HBAl) features were considered to be essential for inhibitory activity in the calling behavior and in vitro pheromone biosynthesis. Active compounds, including yohimbine, mapped well onto all the AR and HBAl features of the hypothesis. Less-active compounds were shown to be unable to achieve an energetically favorable conformation, consistent with our 3D common-feature pharmacophore models. The present hypothesis demonstrates that calling behavior and PBAN-stimulated incorporation of radioactivity are inhibited by tyraminergic antagonists.

Comparative receptor surface analysis of octopaminergic antagonists for the locust neuronal octopamine receptor.

In drug discovery, it is common to have measured activity data for a set of compounds acting upon a particular protein but not to have knowledge of the three-dimensional structure of the protein active site. In the absence of such three-dimensional information, one can attempt to build a hypothetical model of the receptor site that can provide insight about receptor site characteristics. Such a model is known as a comparative receptor surface analysis (CoRSA) model, which provides compact and quantitative descriptors which capture three-dimensional information about a putative receptor site. The quantitative structure-activity relationship (QSAR) of a set of 20 antagonists for octopamine (OA) receptor 3 in locust nervous tissue, was analyzed using CoRSA. Three-dimensional energetics descriptors were calculated from receptor surface model (RSM)-ligand interaction and these three-dimensional descriptors were used in QSAR analysis. The predictive character of the QSAR was further assessed using 24 agonists for OA receptor as test molecules. An RSM was generated using some subset of the most active structures and the results provided useful information in the characterization and differentiation of OA receptor.

Three-dimensional molecular-field analyses of octopaminergic agonists for the cockroach neuronal octopamine receptor.

The quantitative structure-activity relationship of a set of 40 octopaminergic agonists against receptor 2 in cockroach nervous tissue, was analyzed using molecular-field analysis (MFA). MFA on the study set of those compounds evaluated effectively the energy between a probe and a molecular model at a series of points defined by a rectangular grid. Contour surfaces for the molecular fields were presented and the results provided useful information in the characterization and differentiation of octopaminergic receptor.

Three-dimensional pharmacophore hypotheses of octopamine/tyramine agonists which inhibit [1-14C]acetate incorporation in Plodia interpunctella.

Three-dimensional pharmacophore hypotheses were built from a set of 36 octopamine (OA)/tyramine (TA) agonists responsible for the inhibition of sex-pheromone production in Plodia interpunctella. Among the ten chemical-featured models generated by a program Catalyst/Hypo, hypotheses including hydrogen-bond acceptor (HBA), hydrogen-bond acceptor aliphatic (HBAl), hydrophobic (Hp), hydrophobic aromatic (HpAr) and hydrophobic aliphatic (HpAl) features were considered to be important and predictive in evaluating OA/TA agonists. Active agonists mapped well onto all the features of the hypothesis such as HBA, HBAl, Hp, HpAr and HpAl features. On the other hand, inactive compounds were shown to be poorly capable of achieving an energetically favorable conformation shared by the active molecules in order to fit the 3-D chemical-feature pharmacophore models. Those hypotheses are considered to be used in designing new leads for hopefully more active compounds. Further research on the comparison of models from the agonists may help elucidate the mechanisms of OA/TA receptor-ligand interactions.

Inhibitors of calling behavior of Plodia interpunctella.

Some octopamine agonists were found to suppress the calling behavior of the stored product Indian meal moth, Plodia interpunctella. Compounds were screened using a calling behavior bioassay using female P. interpunctella. Four active derivatives, with inhibitory activity at the nanomolar range, were identified in order of decreasing activity: 2-(1-phenylethylamino)-2-oxazoline > 2-(2-ethyl,6-methylanilino)oxazolidine > 2-(2-methyl benzylamino)-2-thiazoline > 2-(2,6-diethylanilino)thiazolidine. Three-dimensional pharmacophore hypotheses were built from a set of 15 compounds. Among the ten common-featured models generated by the program Catalyst/HipHop, a hypothesis including a hydrogen-bond acceptor lipid, a hydrophobic aromatic and two hydrophobic aliphatic features was considered to be essential for inhibitory activity in the calling behavior. Active compounds mapped well onto all the hydrogen-bond acceptor lipid, hydrophobic aromatic and hydrophobic aliphatic features of the hypothesis. On the other hand, less active compounds were shown not to achieve the energetically favorable conformation that is found in the active molecules in order to fit the 3D common-feature pharmacophore models. The present studies demonstrate that inhibition of calling behavior is via an octopamine receptor.

Octopaminergic agonists for the cockroach neuronal octopamine receptor.

The compounds 1-(2,6-diethylphenyl)imidazolidine-2-thione and 2-(2,6-diethylphenyl)imidazolidine showed the almost same activity as octopamine in stimulating adenylate cyclase of cockroach thoracic nervous system among 70 octopamine agonists, suggesting that only these compounds are full octopamine agonists and other compounds are partial octopamine agonists. The quantitative structure-activity relationship of a set of 22 octopamine agonists against receptor 2 in cockroach nervous tissue, was analyzed using receptor surface modeling. Three-dimensional energetics descriptors were calculated from receptor surface model/ligand interaction and these three-dimensional descriptors were used in quantitative structure-activity relationship analysis. A receptor surface model was generated using some subset of the most active structures and the results provided useful information in the characterization and differentiation of octopaminergic receptor.

Three-dimensional common-feature hypotheses of inhibitors of calling behaviour and in vitro [14C]acetate incorporation by pheromone glands of Plodia interpunctella.

Some octopamine (OA) agonists were found to suppress the calling behaviour and pheromone biosynthesis in vitro of the Indian meal moth, Plodia interpunctella (Hübner), a stored-product pest. Compounds were screened using a calling behaviour bioassay of female P interpunctella. Three active derivatives, with activity at the nanomolar level, were identified. In order of decreasing pheromonostatic activity these were: 2-(2-ethyl-6-methylanilino)oxazolidine > 2-(2,6-diethylanilino)thiazolidine > 2-(2,6-diethylanilino)oxazolidine. These compounds showed also in vitro inhibitory activities in de novo pheromone biosynthesis. Three-dimensional pharmacophore hypotheses were built from a set of 19 compounds. Among the ten common-featured models generated by the program Catalyst/HipHop, a hypothesis including a ring aromatic group (RA), a positive ionizable group (PI) and two hydrophobic aliphatic (HpA1) features was considered to be essential for inhibitory activity in the calling behaviour and pheromone biosynthesis in vitro. Active compounds mapped well onto all the RA, PI and HpA1 features of the hypothesis. Less-active compounds were shown not to achieve the energetically favourable conformation which was found in the active molecules in order to fit the 3-D common-feature pharmacophore models. The present studies demonstrate that inhibition of calling behaviour and PBAN-stimulated incorporation of radioactivity is by OA-agonistic activity.

Three-dimensional common-feature hypotheses for octopamine agonist 2-(arylimino)imidazolidines.

Three-dimensional pharmacophore hypotheses were built from a set of 10 octopamine (OA) agonist 2-(Arylimino)imidazolidines (AIIs), 2-(Arylimino)thiazolidines (AITs) and 2-(Arylimino)oxazolidines (AIOs). Among the 10 common-featured models generated by program Catalyst/HipHop, a hypothesis including a ring aromatic (RA), a positive ionizable (PI) and three hydrophobic aliphatic (HpAl) features was considered to be important in evaluating the OA-agonist activity. Active OA agonist 2,6-Et2 AII mapped well onto all the RA, PI and HpAl features of the hypothesis. On the other hand, less active compounds were shown to be difficult to achieve the energetically favorable conformation which is found in the active molecules in order to fit the 3-D common-feature pharmacophore models. Taken together, 2,6-Et2-Ph and foramidine structures are important as OA agonists. The present studies on OA agonists demonstrate that a RA, a PI and three HpAl sites located on the molecule seem to be essential for OA-agonist activity.