Research on marine actinobacteria in India

Marine actinobacteriology is one of the major emerging areas of research in tropics. Marine actinobacteria occur on the sediments and in water and also other biomass (mangrove) and substrates (animal). These organisms are gaining importance not only for their taxonomic and ecological perspectives, but also for their unique metabolites and enzymes. Many earlier studies on these organisms were confined only to the temperate regions. In tropical environment, investigations on them have gained importance only in the last two decades. So far, from the Indian peninsula, 41 species of actinobacteria belonging to 8 genera have been recorded. The genus, Streptomyces of marine origin has been more frequently recorded. Of 9 maritime states of India, only 4 have been extensively covered for the study of marine actinobacteria. Most of the studies conducted pertain to isolation, identification and maintenance of these organisms in different culture media. Further, attention has been focused on studying their antagonistic properties against different pathogens. Their biotechnological potentials are yet to be fully explored.     

An Extracellullar Nuclease from Bacillus Firmus VKPACU-1: Specificity and Mode of Action

An extracellular nuclease from Bacillus firmus VKPACU-1 was multifunctional enzyme, this nuclease hydrolyzed poly U rapidly and more preferentially than the other homopolyribonucleotides. Hydrolysis of RNA this enzyme released mononucleotides in the order 5′UMP > 5′AMP > 5′GMP where as in hydrolysis of DNA the mononucleotides in the order of 5′dAMP > 5′dGMP > 5′dTMP and oligonucleotides. Uridylic linkages in RNA and adenylic linkages in DNA were preferentially cleaved by the nuclease. Nuclease produced oligonucleotides having only 3’ hydroxyl and 5’ phosphate termini. Present nuclease hydrolyzed RNA and DNA released oligonucleotides as major end products and mononucleotides, suggesting an endo mode of action.     

A Dynamin Homolog Promotes the Transition from Hemifusion to Content Mixing in Intracellular Membrane Fusion

The convergence of the antagonistic reactions of membrane fusion and fission at the hemifusion/hemifission intermediate has generated a captivating enigma of whether Soluble N‐ethylmaleimide sensitive factor Attachment Protein Receptor (SNAREs) and dynamin have unusual counter‐functions in fission and fusion, respectively. SNARE‐mediated fusion and dynamin‐driven fission are fundamental membrane flux reactions known to occur during ubiquitous cellular communication events such as exocytosis, endocytosis and vesicle transport. Here we demonstrate the influence of the dynamin homolog Vps1 (Vacuolar protein sorting 1) on lipid mixing and content mixing properties of yeast vacuoles, and on the incorporation of SNAREs into fusogenic complexes. We propose a novel concept that Vps1, through its oligomerization and SNARE domain binding, promotes the hemifusion‐content mixing transition in yeast vacuole fusion by increasing the number of trans‐SNAREs .      

Evidence for the induction of apoptosis by endosulfan in a human T-cell leukemic line

Several organochlorinated pesticides including DDT, PCBs and dieldrin have been reported to cause immune suppression and increase susceptibility to infection in animals. Often this manifestation is accompanied by atrophy of major lymphoid organs. It has been suggested that increased apoptotic cell death leading to altered T-B cell ratios, and loss of regulatory cells in critical numbers leads to perturbations in immune function. The major objective of our study was to define the mechanism by which endosulfan, an organochlorinated pesticide, induces human T-cell death using Jurkat, a human T-cell leukemic cell line, as an in vitro model. We exposed Jurkat cells to varying concentrations of endosulfan for 0-48 h and analyzed biochemical and molecular features characteristic of T-cell apoptosis. Endosulfan lowered cell viability and inhibited cell growth in a dose- and time-dependent manner. DAPI staining was used to enumerate apoptotic cells and we observed that endosulfan at 10-200 M induced a significant percentage of cells to undergo apoptotic cell death. At 48 h, more than 90% cells were apoptotic with 50 M of endosulfan. We confirmed these observations using both DNA fragmentation and annexin-V binding assays. It is now widely being accepted that mitochondria undergo major changes early during the apoptotic process. We examined mitochondrial transmembrane potential (_m) in endosulfan treated cells to understand the role of the mitochondria in T-cell apoptosis. Within 30 min of chemical exposure, a significant percentage of cells exhibited a decreased incorporation of DiOC₆(3), a cationic lipophilic dye into mitochondria indicating the disruption of _m. This drop in _m was both dose- and time-dependent and correlated well with other parameters of apoptosis. We also examined whether this occurred by the down regulation of bcl-2 protein expression that is likely to increase the susceptibility of Jurkat cells to endosulfan toxicity. Paradoxically, the intracellular expression of bcl-2 protein was elevated in a dose dependent manner suggesting endosulfan-induced apoptosis occurred by a non-bcl-2 pathway. Based on these data, as well as those reported elsewhere, we propose the following sequence of events to account for T-cell apoptosis induced by endosulfan: uncoupling of oxidative phosphorylation excess ROS production GSH depletion oxidative stress disruption of _m release of cytochrome C and other apoptosis related proteins to cytosol apoptosis. This study reports for the first time that endosulfan can induce apoptosis in a human T-cell leukemic cell line which may have direct relevance to loss of T cells and thymocytes in vivo. Furthermore, our data strongly support a role of mitochondrial dysfunction and oxidative stress in endosulfan toxicity.     

Synthesis of sugar-amino acid-nucleosides as potential glycosyltransferase inhibitors

Sugar-amino acid-nucleosides (SAAN) were synthesized to mimic glycosyl nucleotide donors based on the hypothesis that a basic amino acid may interact with carboxylate groups of the enzyme in a manner similar to the diphosphate metal ion complex. C-Glycoside analogues of the d-galactopyranose or l-arabinofuranose ring systems, and four amino acids (lysine, glutamine, tryptophan, and histidine), were chosen for this study. The targets were synthesized and tested against GlfT2, a galactofuranosyltransferase essential for cell wall galactan biosynthesis in Mycobacterium tuberculosis. The inhibition assay showed that analogues containing histidine and tryptophan are moderate inhibitors of GlfT2.     

Effect of vermicompost leachate in <Emphasis Type="Italic">Ceratotheca triloba</Emphasis> under nutrient deficiency

Ceratotheca triloba (Bernh.) Hook.f. commonly known as an African foxglove is an indigenous plant which occurs in most parts of South Africa. The species is commonly consumed as a leafy vegetable and utilized for its medicinal properties. Although the high nutritional value of the species and medicinal properties are well documented, information related to critical aspect of cultivation is currently limited. Therefore, this study aimed to evaluate the effect of vermicompost leachate (VCL) on growth, nutritional, phytochemical, and antioxidant levels in C. triloba at different growth stages under nutrient-deficient conditions. After in vitro germination, seedlings were grown in the greenhouse for 2 and 4 months under nitrogen (–N); phosphorus (–P); and potassium (–K) deficiency conditions, and were treated with VCL. Vermicompost leachate did not improve the growth of C. triloba plants under the nutrient-deficient conditions. Although –N-deficient plants with or without VCL caused a decline in growth parameters, they significantly enhanced phytochemicals in 2-month-old plants. In most cases, the application of VCL to –P- and –K-deficient plants improved the photosynthetic pigments, protein, and phenolic, as well as flavonoid accumulation. Harvesting time was also found to play a crucial role in the accumulation of evaluated parameters in nutrient-deprived plants. From these findings, it can be deduced that VCL has a potential to minimize the effect of nutrient deficiency especially under –P and –K deficiency in C. triloba plants.     

Reversibility of oxygen induced inactivation of nitrogenase in some enterobacteria

Aerobic and microaerobic diazotrophs possess numerous oxygen restriction strategies to protect nitrogenase from inactivation by oxygen without interfering with energy generation through oxidative phosphorylation. Protection by conformational change in nitrogenase was first detected and described in Azotobacter. This strategy once considerd unique for Azotobacter has been shown in this study to occur in Citrobacterfreundii (Braak) Werkman and Gillen and Klebsiella pneumoniae subspecies rhinoscleromatis (Trevisan) Migula also. However, in these enteric bacteria the entire enzyme is not protected probably due to the absence of any respiratory protection similar to that found in the aerobe, Azotobacter.