A rapid identification system for metallothionein proteins using expert system

Metallothioneins (MT) are low molecular weight proteins mostly rich in cysteine residues with high metal content. Generally, MT proteins are responsible for regulating the intracellular supply of biologically essential metal ions and they protect cells from the deleterious effects of non-essential polarizable transition and post-transition metal ions. Due to their biological importance, proper characterization of MT is necessary. Here we describe a computer program (ID3 algorithm, a part of Artificial Intelligence) developed using available data for the rapid identification of MT. Tissue samples contains several low molecular weight proteins with different physical, chemical and biological characteristics. The described software solution proposes to categorize MT proteins without aromatic amino acids and high metal content. The proposed solution can be expanded to other types of proteins with specific known characteristics.     

Modeling analysis of GST (glutathione-S-transferases) from Wuchereria bancrofti and Brugia malayi

GST (glutathione S-transferases) are a family of detoxification enzymes that catalyze the conjugation of reduced GSH (glutathione) to xenobiotic (endogenous electrophilic) compounds. GST from Wb (Wuchereria bancrofti) and Bm (Brugia malayi) are significantly different from human GST in sequence and structure. Thus, Wb-GST and Bm-GST are potential chemotherapeutic targets for anti-filarial treatment. Comparison of modeled Wb and Bm GST with human GST show structural difference between them. Analysis of the active site residues for the binding of electrophilic co-substrates provides insight towards the design of parasite specific GST inhibitors.     

Curcumin Modulates the Inflammatory Response and Inhibits Subsequent Fibrosis in a Mouse Model of Viral-induced Acute Respiratory Distress Syndrome

Acute Respiratory Distress Syndrome (ARDS) is a clinical syndrome characterized by diffuse alveolar damage usually secondary to an intense host inflammatory response of the lung to a pulmonary or extrapulmonary infectious or non-infectious insult often leading to the development of intra-alveolar and interstitial fibrosis. Curcumin, the principal curcumoid of the popular Indian spice turmeric, has been demonstrated as an anti-oxidant and anti-inflammatory agent in a broad spectrum of diseases. Using our well-established model of reovirus 1/L-induced acute viral pneumonia, which displays many of the characteristics of the human ALI/ARDS, we evaluated the anti-inflammatory and anti-fibrotic effects of curcumin. Female CBA/J mice were treated with curcumin (50 mg/kg) 5 days prior to intranasal inoculation with 10⁷ pfu reovirus 1/L and daily, thereafter. Mice were evaluated for key features associated with ALI/ARDS. Administration of curcumin significantly modulated inflammation and fibrosis, as revealed by histological and biochemical analysis. The expression of IL-6, IL-10, IFNγ, and MCP-1, key chemokines/cytokines implicated in the development of ALI/ARDS, from both the inflammatory infiltrate and whole lung tissue were modulated by curcumin potentially through a reduction in the phosphorylated form of NFκB p65. While the expression of TGFß1 was not modulated by curcumin, TGFß Receptor II, which is required for TGFß signaling, was significantly reduced. In addition, curcumin also significantly inhibited the expression of α-smooth muscle actin and Tenascin-C, key markers of myofibroblast activation. This data strongly supports a role for curcumin in modulating the pathogenesis of viral-induced ALI/ARDS in a pre-clinical model potentially manifested through the alteration of inflammation and myofibroblast differentiation.     

Gateway vectors for the production of combinatorially-tagged His6-MBP fusion proteins in the cytoplasm and periplasm of Escherichia coli

Many proteins that accumulate in the form of insoluble aggregates when they are overproduced in Escherichia coli can be rendered soluble by fusing them to E. coli maltose binding protein (MBP), and this will often enable them to fold in to their biologically active conformations. Yet, although it is an excellent solubility enhancer, MBP is not a particularly good affinity tag for protein purification. To compensate for this shortcoming, we have engineered and successfully tested Gateway destination vectors for the production of dual His6MBP-tagged fusion proteins in the cytoplasm and periplasm of E. coli. The MBP moiety improves the yield and solubility of its fusion partners while the hexahistidine tag (His-tag) serves to facilitate their purification. The availability of a vector that targets His6MBP fusion proteins to the periplasm expands the utility of this dual tagging approach to include proteins that contain disulfide bonds or are toxic in the bacterial cytoplasm.     

Management of filariasis using prediction rules derived from data mining

The present paper demonstrates the application of CART (classification and regression trees) to control a mosquito vector (Culex quinquefasciatus) for bancroftian filariasis in India. The database on filariasis and a commercially available software CART (Salford systems Inc. USA) were used in this study. Baseline entomological data related to bancroftian filariasis was utilized for deriving prediction rules. The data was categorized into three different aspects, namely (1) mosquito abundance, (2) meteorological and (3) socio-economic details. This data was taken from a database developed for a project entitled "Database management system for the control of bancroftian filariasis" sponsored by Ministry of Communication and Information Technology (MC&IT), Government of India, New Delhi. Predictor variables (maximum temperature, minimum temperature, rain fall, relative humidity, wind speed, house type) were ranked by CART according to their influence on the target variable (month). The approach is useful for forecasting vector (mosquito) densities in forthcoming seasons.     

A web based relational database management system for filariasis control

The present study describes a RDBMS (relational database management system) for the effective management of Filariasis, a vector borne disease. Filariasis infects 120 million people from 83 countries. The possible re-emergence of the disease and the complexity of existing control programs warrant the development of new strategies. A database containing comprehensive data associated with filariasis finds utility in disease control. We have developed a database containing information on the socio-economic status of patients, mosquito collection procedures, mosquito dissection data, filariasis survey report and mass blood data. The database can be searched using a user friendly web interface.

Availability     

New Bacillus thuringiensis strain isolated from the gut of Malabari goat is effective against Tetranychus macfarlanei

This study illustrates a novel strain (designated as BPU5) of Bacillus thuringiensis (Bt) isolated from the rumen of Malabari goat, capable of producing polymorphic δ‐endotoxin crystals concomitantly with sporulation in Luria–Bertani medium (LB), and the δ‐endotoxin was efficient to combat Tetranychus macfarlanei, a devastating mite. Polymorphic δ‐endotoxin crystals produced were assessed by scanning electron microscopy and monitored its production concomitantly with sporulation in LB with or without sugar supplements. Toxicity of the δ‐endotoxin was assessed on T. macfarlanei using leaf disc bioassay method. Mortality rate was determined by comparing the survival of mites on the diet (prepared in 10% sucrose and powdered rice husk) containing different concentrations (1–10 mg/ml) of 72‐h‐old crude pellet (dried mixture of δ‐endotoxin (17 mg/g pellet), endospores and a few vegetative cells) or control diet with autoclaved pellet. The maximum production (1.39 mg/ml) of δ‐endotoxin was observed at 72 h in LB. Among the sugars (glucose, sucrose, maltose or lactose) tested as additional carbon source, glucose (8 g/l) enhanced (1.82 mg/ml) the production of δ‐endotoxin by 30%. The lethal concentration (LC₅₀) required to kill 50% mites was estimated as 8.024 mg/ml. The δ‐endotoxin produced by B. thuringiensis BPU5 is shown to efficiently combat T. macfarlanei, a devastating mite infesting agricultural fields.     

Repeat-Associated Fission Yeast-Like Regional Centromeres in the Ascomycetous Budding Yeast Candida tropicalis

The centromere, on which kinetochore proteins assemble, ensures precise chromosome segregation. Centromeres are largely specified by the histone H3 variant CENP-A (also known as Cse4 in yeasts). Structurally, centromere DNA sequences are highly diverse in nature. However, the evolutionary consequence of these structural diversities on de novo CENP-A chromatin formation remains elusive. Here, we report the identification of centromeres, as the binding sites of four evolutionarily conserved kinetochore proteins, in the human pathogenic budding yeast Candida tropicalis. Each of the seven centromeres comprises a 2 to 5 kb non-repetitive mid core flanked by 2 to 5 kb inverted repeats. The repeat-associated centromeres of C. tropicalis all share a high degree of sequence conservation with each other and are strikingly diverged from the unique and mostly non-repetitive centromeres of related Candida species—Candida albicans, Candida dubliniensis, and Candida lusitaniae. Using a plasmid-based assay, we further demonstrate that pericentric inverted repeats and the underlying DNA sequence provide a structural determinant in CENP-A recruitment in C. tropicalis, as opposed to epigenetically regulated CENP-A loading at centromeres in C. albicans. Thus, the centromere structure and its influence on de novo CENP-A recruitment has been significantly rewired in closely related Candida species. Strikingly, the centromere structural properties along with role of pericentric repeats in de novo CENP-A loading in C. tropicalis are more reminiscent to those of the distantly related fission yeast Schizosaccharomyces pombe. Taken together, we demonstrate, for the first time, fission yeast-like repeat-associated centromeres in an ascomycetous budding yeast.     

Solubility-enhancing proteins MBP and NusA play a passive role in the folding of their fusion partners

It is well established that certain highly soluble proteins have the ability to enhance the solubility of their fusion partners. However, very little is known about how different solubility enhancers compare in terms of their ability to promote the proper folding of their passenger proteins. We compared the ability of two well-known solubility enhancers, Escherichia coli maltose-binding protein (MBP) and N utilization substance A (NusA), to improve the solubility and promote the proper folding of a variety of passenger proteins that are difficult to solubilize. We used an intracellular processing system to monitor the solubility of these passenger proteins after they were cleaved from MBP and NusA by tobacco etch virus protease. In addition, the biological activity of some fusion proteins was compared to serve as a more quantitative indicator of native structure. The results indicate that MBP and NusA have comparable solubility-enhancing properties. Little or no difference was observed either in the solubility of passenger proteins after intracellular processing of the MBP and NusA fusion proteins or in the biological activity of solubilized passenger proteins, suggesting that the underlying mechanism of solubility enhancement is likely to be similar for both the proteins, and that they play a passive role rather than an active one in the folding of their fusion partners.     

Induction of sister chromatid exchanges in traffic policemen exposed to vehicular exhaust

In urban areas there is an explosive growth of population and the number of automobiles. The ever-increasing vehicular traffic density is posing continued threat to the ambient air quality. Traffic policemen as a group of workers are exposed occupationally to the pollutants from vehicular exhaust. Sister chromatid exchanges (SCEs) as a biomarker of the pollutant's effect, were analyzed in peripheral blood lymphocytes of 85 traffic policemen and 60 control subjects. There was a significant increase in the mean SCEs+/-S.D./cell in the exposed group (9.31+/-5.29) when compared to the controls (4.18+/-1.85). Thus the present study concludes that vehicular exhaust might induce cytogenetic damage in traffic police. Further, the more pronounced frequency of SCEs observed in the smoking traffic policemen than in the non-smoking group suggests the joint effect of smoking and vehicular exhaust.