In vitro selection of endosulfan-tolerant strains of Brassica compestris (cv. Brown Sarson)

Endosulfan tolerant lines of mustard (Brassica campestris cv. Brown Sarson) have been developed through tissue culture methods. Cotyledonary expiants excised from eight day old in vitro grown seedlings were used for inducing callus. Fast growing friable callus was then transferred to MS medium containing (0.1–2.0 ugl^–1) endosulfan for selection. Five alternating exposures with and without endosulfan containing medium yielded an endosulfan tolerant cell line (ETL). The plants regenerated from ETL were found to tolerate three fold higher concentrations of endosulfan. Callus induced from randomly selected endosulfan tolerant regenerated plants were also tolerant to 3.0 ugl endosulfan, thereby, suggesting that tolerance has been acquired at the gene level.Biochemical investigation revealed higher levels of total free sugar, free amino acids, protein and activity of peroxidase in the tolerant cell line.Abbreviations MS Murashige and Skoog medium (1962) - NSM non-selective medium - SM selective medium - BAP 6-Benzylaminopurine - NAA -naphthaleneacetic acid - Z zeatin     

Protective effect of coenzyme Q10 in simvastatin and gemfibrozil induced rhabdomyolysis in rats

Administration of simvastatin (80 mg/kg, po. evening dose) and gemfibrozil (600 mg/kg, po twice) for 30 days produced significant decrease in the level of reduced glutathione, superoxide dismutase, catalase and increase in the level of lipid peroxidation and various serum parameters (creatine phosphokinase, lactate dehydrogenase, serum glutamate oxaloacetate transaminase, creatinine, urea and blood urea nitrogen). This suggested involvement of oxidative stress in rhabdomyolysis. Increase in the level of reduced glutathione, superoxide dismutase, catalase and decrease in the level of lipid peroxidation and serum parameters after administration of antioxidant CoQ10 (10 mg/kg.ip) proved the protective effect of CoQ10 in rhabdomyolysis.     

Autophagy in hypothalamic AgRP neurons regulates food intake and energy balance

Macroautophagy is a lysosomal degradative pathway that maintains cellular homeostasis by turning over cellular components. Here we demonstrate a role for autophagy in hypothalamic agouti-related peptide (AgRP) neurons in the regulation of food intake and energy balance. We show that starvation-induced hypothalamic autophagy mobilizes neuron-intrinsic lipids to generate endogenous free fatty acids, which in turn regulate AgRP levels. The functional consequences of inhibiting autophagy are the failure to upregulate AgRP in response to starvation, and constitutive increases in hypothalamic levels of pro-opiomelanocortin and its cleavage product α-melanocyte-stimulating hormone that typically contribute to a lean phenotype. We propose a conceptual framework for considering how autophagy-regulated lipid metabolism within hypothalamic neurons may modulate neuropeptide levels to have immediate effects on food intake, as well as long-term effects on energy homeostasis. Regulation of hypothalamic autophagy could become an effective intervention in conditions such as obesity and the metabolic syndrome.     

Racemic Resolution of some <Emphasis Type="SmallCaps">dl</Emphasis>-Amino Acids using <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis><Emphasis Type="SmallCaps">l</Emphasis>-Amino Acid Oxidase

The ability of Aspergillus fumigatus l-amino acid oxidase (l-aao) to cause the resolution of racemic mixtures of dl-amino acids was investigated with dl-alanine, dl-phenylalanine, dl-tyrosine, and dl-aspartic acid. A chiral column, Crownpak CR+ was used for the analysis of the amino acids. The enzyme was able to cause the resolution of the three dl-amino acids resulting in the production of optically pure d-alanine (100% resolution), d-phenylalanine (80.2%), and d-tyrosine (84.1%), respectively. The optically pure d-amino acids have many uses and thus can be exploited industrially. This is the first report of the use of A. fumigatus l-amino acid oxidase for racemic resolution of dl-amino acids.     

Incorporation of biological knowledge into distance for clustering genes

In this paper we propose a data based algorithm to marry existing biological knowledge (e.g., functional annotations ofgenes) with experimental data (gene expression profiles) in creating an overall dissimilarity that can be used with anyclustering algorithm that uses a general dissimilarity matrix. We explore this idea with two publicly available geneexpression data sets and functional annotations where the results are compared with the clustering results that uses only theexperimental data. Although more elaborate evaluations might be called for, the present paper makes a strong case forutilizing existing biological information in the clustering process.

Availability     

Mapping of the CD23 Binding Site on Immunoglobulin E (IgE) and Allosteric Control of the IgE-Fc{epsilon}RI Interaction

IgE, the antibody that mediates allergic responses, acts as part of a self-regulating protein network. Its unique effector functions are controlled through interactions of its Fc region with two cellular receptors, FcεRI on mast cells and basophils and CD23 on B cells. IgE cross-linked by allergen triggers mast cell activation via FcεRI, whereas IgE-CD23 interactions control IgE expression levels. We have determined the CD23 binding site on IgE, using a combination of NMR chemical shift mapping and site-directed mutagenesis. We show that the CD23 and FcεRI interaction sites are at opposite ends of the Cε3 domain of IgE, but that receptor binding is mutually inhibitory, mediated by an allosteric mechanism. This prevents CD23-mediated cross-linking of IgE bound to FcεRI on mast cells and resulting antigen-independent anaphylaxis. The mutually inhibitory nature of receptor binding provides a degree of autonomy for the individual activities mediated by IgE-FcεRI and IgE-CD23 interactions.     

Fibrillation of hen egg white lysozyme triggers reduction of copper(II)

Copper is known to exert diverse effects on the self-association of proteins and has been found in amyloid deposits that are involved in neurodegenerative disease processes. The effects of the metal ion on the protein during fibrillation were investigated by fluorescence, circular dichroism spectroscopy and fluorescence microscopy. We report for the first time, the complete reduction of Cu(II)→Cu(I) in vitro during fibrillation of hen egg white lysozyme at pH 7. This was confirmed by the lack of any signal for Cu(II) in electron paramagnetic resonance spectroscopy and quantification of Cu(I) was achieved by a bathocuproine disulfonate based assay.     

Single mutation at P1 of a chymotrypsin inhibitor changes it to a trypsin inhibitor: X-ray structural (2.15 Å) and biochemical basis

Change in specificity, caused by the mutations at P1 site, of the serine protease inhibitors of different families is reported in the literature, but Kunitz (STI) family inhibitors are almost unexplored in this regard. In this paper, we present the crystal structure of a P1 variant of winged bean chymotrypsin inhibitor (WCI) belonging to Kunitz (STI) family, supplemented by biochemical, phylogenetic and docking studies on the mutant. A single mutation (Leu → Arg) at P1 converted WCI to a strong inhibitor of trypsin with an association constant of 4.8 × 10¹⁰ M^?1 which is comparable to other potent trypsin inhibitors of the family. The crystal structure (2.15 Å) of this mutant (L65R) shows that its reactive site loop conformation deviates from that of WCI and adopts a structure similar to that of Erythrina caffra trypsin inhibitor (ETI) belonging to the same family. Mutation induced structural changes have also been propagated in a concerted manner to the neighboring conserved scaffolding residue Asn14, such that the side chain of this residue took an orientation similar to that of ETI and optimized the hydrogen bonds with the loop residues. While docking studies provide information about the accommodation of non-specific residues in the active site groove of trypsin, the basis of the directional alteration of the reactive site loop conformation has been understood through sequence analysis and related phylogenetic studies.