Double drugging of prolyl-tRNA synthetase provides a new paradigm for anti-infective drug development

Toxoplasmosis is caused by Toxoplasma gondii and in immunocompromised patients it may lead to seizures, encephalitis or death. The conserved enzyme prolyl-tRNA synthetase (PRS) is a validated druggable target in Toxoplasma gondii but the traditional ‘single target–single drug’ approach has its caveats. Here, we describe two potent inhibitors namely halofuginone (HFG) and a novel ATP mimetic (L95) that bind to Toxoplasma gondii PRS simultaneously at different neighbouring sites to cover all three of the enzyme substrate subsites. HFG and L95 act as one triple-site inhibitor in tandem and form an unusual ternary complex wherein HFG occupies the 3’-end of tRNA and the L-proline (L-pro) binding sites while L95 occupies the ATP pocket. These inhibitors exhibit nanomolar IC₅₀ and EC₅₀ values independently, and when given together reveal an additive mode of action in parasite inhibition assays. This work validates a novel approach and lays a structural framework for further drug development based on simultaneous targeting of multiple pockets to inhibit druggable proteins.     

Designing an immunoinformatic vaccine for peri-implantitis using a structural biology approach

ObjectivesPeri-implantitis is a destructive inflammatory process that affects the soft and hard tissues around dental implants. porphyromonas gingivalis, an anaerobic gram-negative bacterium, appears to be the main culprit. Since there is no efficient and specific vaccine to treat peri-implantitis, the goal of our research has been to develop a multi-epitope vaccination utilizing an immunoinformatics approach that targeted P. gingivalis type I fim A.Materials and methodsP. gingivalis peptides 6JKZ and 6KMF are suitable for vaccine development. B- and T-cell epitopes from 6KMF and 6JKZ were detected and evaluated based on critical factors to produce a multi-epitope vaccine construct. It was assessed based on allergenicity, antigenicity, stability. The vaccine's dual major histocompatibility complex (MHC-I and MHC-II) binding epitopes allowed it to reach a larger population. P. gingivalis fimbriae induce immune subversion through TLR -CXCR4 receptor complex pathway. The ClusPro 2.0 server was used to do the molecular docking using TLR2 - CXCR4 and vaccine epitopes as receptor and ligand respectively.ResultsThe designed vaccine was non-allergenic and had a high antigenicity, solubility, and stability. The 3D structure of the vaccine revealed strong interaction with CXCR4(TLR2) using molecular docking. The vaccine-CXCR4 interface was more consistent, possibly because the vaccination has a higher affinity for the CXCR4-TLR2 complex.ConclusionThis study details the vaccine's distinct and sustained interaction with the CXCR4(TLR2) immunological receptor and its consistent and effective utterance in the bacterial system. As a result, our vaccine formulation will evoke a significant memory response and induce an adaptive immune response against P. gingivalis.     

Molecular docking analysis of Clostridium perfringens beta toxin model with potential inhibitors from the ZINC database

Beta toxin from Clostridium perfringens after being secreted in gut is capable of causing necrotic enteritis in humans and several other animal species and does not respond to routinely used antibiotics. Therefore, there is a need to design an effective inhibitor for the Clostridium perfringens beta toxin (CPB) using cutting edge drug discovery technologies. Hence, potential CPB inhibitors were identified using computer aided screening of compounds from the ZINC database. Further, we document the molecular docking analysis of Clostridium perfringens beta toxin model (that revealed 4 binding pockets, A-D) with the identified potential inhibitors. We show that ZINC291192 [N-[(1-methylindol-3-yl) methyl eneamino]-7,10-dioxabicyclo[4.4.0]deca-2,4,11-triene-8- carboxamide] has optimal binding features with calculated binding energy of -10.38 kcal/mol and inhibition constant of 24.76 nM for further consideration.     

Evidence for photosynthetic independence of viral multiplication in cyanophage LPP-1 infected cyanobacterium Phormidium uncinatum

Abstract Pigment decomposition, oxygen evolution and CO₂ fixation were measured in the cyanobacterium Phormidium uncinatum after infection with cyanophage LPP-1, under light and dark conditions. A gradual decrease in para benzoquinone supported O₂ evolution, chlorophyll a and phycocyanin level were noticed after 6 h of infection. These results demonstrated decreased photosynthetic activity of the host P. uncinatum prior to the start of LPP-1 multiplication. Metabolic inhibitor investigations confirmed that the cyanophage LPP-1 multiplication was independent of host photosynthesis.     

Fiber optic in vivo imaging in the mammalian nervous system

The compact size, mechanical flexibility, and growing functionality of optical fiber and fiber optic devices are enabling several new modalities for imaging the mammalian nervous system in vivo. Fluorescence microendoscopy is a minimally invasive fiber modality that provides cellular resolution in deep brain areas. Diffuse optical tomography is a non-invasive modality that uses assemblies of fiber optic emitters and detectors on the cranium for volumetric imaging of brain activation. Optical coherence tomography is a sensitive interferometric imaging technique that can be implemented in a variety of fiber based formats and that might allow intrinsic optical detection of brain activity at a high resolution. Miniaturized fiber optic microscopy permits cellular level imaging in the brains of behaving animals. Together, these modalities will enable new uses of imaging in the intact nervous system for both research and clinical applications.     

Identifying patterns of DNA for tumor diagnosis using capillary electrophoresis-amplified fragment length polymorphism (CE-AFLP) screening

Amplified Fragment Length Polymorphism (AFLP) screening is a genome-wide genotyping strategy that has been widely used in plants and bacteria, but little has been reported concerning its use in humans. We investigated if the AFLP procedure could be coupled with high-throughput capillary electrophoresis (CE) for use in tumor diagnosis and classification. Using CE-AFLP, a series of molecular 'fingerprints' were generated for a set of gastric tumor and normal genomic DNA samples. The CE-AFLP procedure was qualitatively and quantitatively robust, and a variety of clustering tools were used to identify a specific DNA marker 'pattern' of 20 features that classified the tumor and normal samples to reasonable degrees of accuracy (Sensitivity 95%, Specificity 80%). The CE-AFLP-based approach also correctly classified 16 tumor samples, which in a previous study had exhibited no detectable genomic aberrations by comparative genome hybridization (CGH). This is the first reported application of CE-AFLP screening in tumor diagnosis. As the procedure is relatively inexpensive and requires minimal prior sequence knowledge and biological material, we suggest that CE-AFLP-based protocols may represent a promising new approach for DNA-based cancer screening and diagnosis.     

Predictors of mortality of patients with acute respiratory failure secondary to chronic obstructive pulmonary disease admitted to an intensive care unit: A one year study

Background

Patients with acute exacerbation of chronic obstructive pulmonary disease (COPD) commonly require hospitalization and admission to intensive care unit (ICU). It is useful to identify patients at the time of admission who are likely to have poor outcome. This study was carried out to define the predictors of mortality in patients with acute exacerbation of COPD and to device a scoring system using the baseline physiological variables for prognosticating these patients.

Methods

Eighty-two patients with acute respiratory failure secondary to COPD admitted to medical ICU over a one-year period were included. Clinical and demographic profile at the time of admission to ICU including APACHE II score and Glasgow coma scale were recorded at the time of admission to ICU. In addition, acid base disorders, renal functions, liver functions and serum albumin, were recorded at the time of presentation. Primary outcome measure was hospital mortality.

Results

Invasive ventilation was required in 69 patients (84.1%). Fifty-two patients survived to hospital discharge (63.4%). APACHE II score at the time of admission to ICU {odds ratio (95 % CI): 1.32 (1.138–1.532); p < 0.001} and serum albumin (done within 24 hours of admission) {odds ratio (95 % CI): 0.114 (0.03-0.432); p = 0.001}. An equation, constructed using the adjusted odds ratio for the two parameters, had an area under the ROC curve of 91.3%. For the choice of cut-off, sensitivity, specificity, positive and negative predictive value for predicting outcome was 90%, 86.5%, 79.4% and 93.7%.

Conclusion

APACHE II score at admission and SA levels with in 24 hrs after admission are independent predictors of mortality for patients with COPD admitted to ICU. The equation derived from these two parameters is useful for predicting outcome of these patients.     

Melatonin blocks inhibitory effects of prolactin on photoperiodic induction of gain in body mass,testicular growth and feather regeneration in the migratory male redheaded bunting (<Emphasis Type="Italic">Emberiza bruniceps</Emphasis>)

Little is known about how hormones interact in the photoperiodic induction of seasonal responses in birds. In this study, two experiments determined if the treatment with melatonin altered inhibitory effects of prolactin on photoperiodic induction of seasonal responses in the Palearctic-Indian migratory male redheaded bunting Emberiza bruniceps. Each experiment employed three groups (N = 6–7 each) of photosensitive birds that were held under 8 hours light: 16 hours darkness (8L:16D) since early March. In the experiment 1, beginning in mid June 2001, birds were exposed to natural day lengths (NDL) at 27 degree North (day length = ca.13.8 h, sunrise to sunset) for 23 days. In the experiment 2, beginning in early April 2002, birds were exposed to 14L:10D for 22 days. Beginning on day 4 of NDL or day 1 of 14L:10D, they received 10 (experiment 1) or 13 (experiment 2) daily injections of both melatonin and prolactin (group 1) or prolactin alone (group 2) at a dose of 20 microgram per bird per day in 200 microliter of vehicle. Controls (group 3) received similar volume of vehicle. Thereafter, birds were left uninjected for the next 10 (experiment 1) or 9 days (experiment 2). All injections except those of melatonin were made at the zeitgeber time 10 (ZT 0 = time of sunrise, experiment 1; time of lights on, experiment 2); melatonin was injected at ZT 9.5 and thus 0.5 h before prolactin. Observations were recorded on changes in body mass, testicular growth and feather regeneration.