Synthesis of glucose-templated lysine analogs and incorporation into the antimicrobial dipeptide sequence kW-OBn

The synthesis of two glucose-templated (GlcT) lysine analogs GlcTK and GlcTk in which the side chain of D- and L-lysine (k and K) is conformationally constrained via incorporation into a D-glucose scaffold is described. A key-step in the synthesis is a high yielding, reductive ring opening of an exocyclic glucose-derived epoxide to form a alpha-hydroxy ester that can be converted into GlcTK and GlcTk. To demonstrate the use of these building blocks in peptide synthesis, we replaced D-lysine in the antimicrobial dipeptide sequence kW-OBn (W=L-tryptophan) and determined the antibacterial activity against various gram-positive and gram-negative organisms. Our results show that the replacement of D-lysine by unprotected GlcTk in dipeptide kW-OBn results in reduced antibacterial activity.     

Improved activity of enzymes in mixed cationic reverse micelles with imidazolium-based surfactants

This work reports the significant enhancement in performance of interfacially active enzymes, Chromobacterium viscosum (CV) lipase and horseradish peroxidase (HRP) in mixed reverse micelles of cetyltrimethylammonium bromide (CTAB) and imidazolium-based amphiphiles having varying tail lengths. Lipase activity in these mixed systems was always higher than that in the individual cationic reverse micelles of CTAB or any imidazolium surfactant, highest being observed in the mixed system of CTAB (50 mM) and 6 (1-tetradecyl-3-methyl imidazolium bromide, 40 mM)/water/isooctane/n-hexanol (0.24 M), second-order rate constant, k2=1301+/-5 cm3 g(-1)s(-1), approximately 200% higher compared to that in CTAB and approximately 65% more than the most popular AOT-microemulsion. Activity increased with concentration of imidazolium surfactant and also with its alkyl tail length. To have a more profound view on the structure-activity relationship, CTAB was replaced by cetyltriethylammonium bromide (CTEAB) and cetyltripropylammonium bromide (CTPAB) with subsequent increase in the headgroup size. The generalized influence of these mixed cationic systems on surface-active enzyme was also verified using HRP, where the activity improved approximately 100%. This enhancement in enzyme activity is presumably due to the activating effect of the imidazolium cation in the enzymatic reactions by improving the nucleophilicity of interfacial water in vicinity of enzyme through hydrogen bonding.     

Synthesis,characterization, redox behavior,DNA and protein binding and antibacterial activity studies of ruthenium(II) complexes of bidentate schiff bases

Two new ruthenium(II) complexes of Schiff base ligands (L) derived from cinnamaldehyde and ethylenediamine formulated as [Ru(L)(bpy)₂](ClO₄)₂, where L¹ = N,N’-bis(4-nitrocinnamald-ehyde)ethylenediamine and L² = N,N’-bis(2-nitrocinnamaldehyde)-ethylenediamine for complex 1 and 2, respectively, were isolated in pure form. The complexes were characterized by physicochemical and spectroscopic methods. The electrochemical behavior of the complexes showed the Ru(III)/Ru(II) couple at different potentials with quasi-reversible voltammograms. The interaction of the complexes with calf thymus DNA (CT-DNA) using absorption, emission spectral studies and electrochemical techniques have been used to determine the binding constant, K_b and the linear Stern–Volmer quenching constant, K_SV. The results indicate that the ruthenium(II) complexes interact with CT-DNA strongly in a groove binding mode. The interactions of bovine serum albumin (BSA) with the complexes were also investigated with the help of absorption and fluorescence spectroscopy tools. Absorption spectroscopy proved the formation of a ground state BSA-[Ru(L)(bpy)₂](ClO₄)₂ complex. The antibacterial study showed that the Ru(II) complexes (1 and 2) have better activity than the standard antibiotics but weak activity than the ligands.     

Quantifying surface gradients with a 2-band Enhanced Vegetation Index (EVI2)

Quantification of spatial and temporal heterogeneity has been given much attention in order to link ecological patterns to processes. The patch mosaic model, as an operational paradigm, has led to major advances in the field of quantitative landscape ecology. However, it is more realistic to conceptualize landscapes based on continuous rather than discrete spatial heterogeneity. While a conceptual shift has been proposed to supplement the patch mosaic model, few studies use the surface gradient model as a context. This paper explores some comparatively less-utilized metrics to quantify surface gradients in a protected landscape in Central India. Since surface metrics would require continuous variables capable of representing landscape characteristics, this study also explores the utility of a 2-band Enhanced Vegetation Index (EVI2) as a gradient surface. Findings suggest EVI2 relates strongly with discrete land cover classes and thus has potential to describe landscape characteristics without incorporating error through subjectivity. Surface metrics used in this study show potential to be effectively used in landscape level studies. However, these metrics were not developed for landscape level studies and should be used with caution, especially when dealing with multi-scale patterns and processes. Nevertheless, with the rapidly emerging field of surface metrology more studies need to apply these tools to quantify surface gradients and test the robustness of such metrics.     

The affinity concept in bioseparation: evolving paradigms and expanding range of applications

The meaning of the word affinity in the context of protein separation has undergone evolutionary changes over the years. The exploitation of molecular recognition phenomenon is no longer limited to affinity chromatography modes. Affinity based separations today include precipitation, membrane based purification and two-phase/three-phase extractions. Apart from the affinity ligands, which have biological relationship (in vivo) with the target protein, a variety of other ligands are now used in the affinity based separations. These include dyes, chelated metal ions, peptides obtained by phage display technology, combinatorial synthesis, ribosome display methods and by systematic evolution of ligands by exponential enrichment (SELEX). Molecular modeling techniques have also facilitated the designing of biomimetic ligands. Fusion proteins obtained by recombinatorial methods have emerged as a powerful approach in bioseparation. Overexpression in E. coli often result in inactive and insoluble inclusion bodies. A number of interesting approaches are used for simultaneous refolding and purification in such cases. Proteomics also needs affinity chromatography to reduce the complexity of the system before analysis by electrophoresis and mass spectrometry are made. At industrial level, validation, biosafety and process hygiene are also important aspects. This overview looks at these evolving paradigms and various strategies which utilize affinity phenomenon for protein separations.     

Molecular cloning, characterization, and engineering of xylitol dehydrogenase from Debaryomyces hansenii

Because of its natural ability to utilize both xylose and arabinose, the halotolerant and osmotolerant yeast Debaryomyces hansenii is considered as a potential microbial platform for exploiting lignocellulosic biomass. To gain better understanding of the xylose metabolism in D. hansenii, we have cloned and characterized a xylitol dehydrogenase gene (DhXDH). The cloned gene appeared to be essential for xylose metabolism in D. hansenii as the deletion of this gene abolished the growth of the cells on xylose. The expression of DhXDH was strongly upregulated in the presence of xylose. Recombinant DhXdhp was expressed and purified from Escherichia coli. DhXdhp was highly active against xylitol and sorbitol as substrate. Our results showed that DhXdhp was thermo-sensitive, and except this, its biochemical properties were quite comparable with XDH from other yeast species. Furthermore, to make this enzyme suitable for metabolic engineering of D. hansenii, we have improved its thermotolerance and modified cofactor requirement through modelling and mutagenesis approach.     

Central effects of neuromedin U in the regulation of energy homeostasis

Neuromedin U (NMU) is a brain-gut peptide whose peripheral activities are well-understood but whose central actions have yet to be clarified. The recent identification of two NMU receptors in rat brain has provided a springboard for further investigation into its role in the central nervous system. Intracerebroventricular administration of NMU to free-feeding rats decreased food intake and body weight. Conversely, NMU increased gross locomotor activity, body temperature, and heat production. NMU, a potent endogenous anorectic peptide, serves as a catabolic signaling molecule in the brain. Further investigation of the biochemical and physiological functions of NMU will help our better understanding of the mechanisms of energy homeostasis.     

The Vibrio cholerae Extracellular Chitinase ChiA2 Is Important for Survival and Pathogenesis in the Host Intestine

In aquatic environments, Vibrio cholerae colonizes mainly on the chitinous surface of copepods and utilizes chitin as the sole carbon and nitrogen source. Of the two extracellular chitinases essential for chitin utilization, the expression of chiA2 is maximally up-regulated in host intestine. Recent studies indicate that several bacterial chitinases may be involved in host pathogenesis. However, the role of V. cholerae chitinases in host infection is not yet known. In this study, we provide evidence to show that ChiA2 is important for V. cholerae survival in intestine as well as in pathogenesis. We demonstrate that ChiA2 de-glycosylates mucin and releases reducing sugars like GlcNAc and its oligomers. Deglycosylation of mucin corroborated with reduced uptake of alcian blue stain by ChiA2 treated mucin. Next, we show that V. cholerae could utilize mucin as a nutrient source. In comparison to the wild type strain, ΔchiA2 mutant was 60-fold less efficient in growth in mucin supplemented minimal media and was also ∼6-fold less competent to survive when grown in the presence of mucin-secreting human intestinal HT29 epithelial cells. Similar results were also obtained when the strains were infected in mice intestine. Infection with the ΔchiA2 mutant caused ∼50-fold less fluid accumulation in infant mice as well as in rabbit ileal loop compared to the wild type strain. To see if the difference in survival of the ΔchiA2 mutant and wild type V. cholerae was due to reduced adhesion of the mutant, we monitored binding of the strains on HT29 cells. The initial binding of the wild type and mutant strain was similar. Collectively these data suggest that ChiA2 secreted by V. cholerae in the intestine hydrolyzed intestinal mucin to release GlcNAc, and the released sugar is successfully utilized by V. cholerae for growth and survival in the host intestine.     

Somatostatin suppresses ghrelin secretion from the rat stomach

Ghrelin is an acylated peptide that stimulates food intake and the secretion of growth hormone. While ghrelin is predominantly synthesized in a subset of endocrine cells in the oxyntic gland of the human and rat stomach, the mechanism regulating ghrelin secretion remains unknown. Somatostatin, a peptide produced in the gastric oxyntic mucosa, is known to suppress secretion of several gastrointestinal peptides in a paracrine fashion. By double immunohistochemistry, we demonstrated that somatostatin-immunoreactive cells contact ghrelin-immunoreactive cells. A single intravenous injection of somatostatin reduced the systemic plasma concentration of ghrelin in rats. Continuous infusion of somatostatin into the gastric artery of the vascularly perfused rat stomach suppressed ghrelin secretion in both dose- and time-dependent manner. These findings indicate that ghrelin secretion from the stomach is regulated by gastric somatostatin.     

Hydro and halo priming: influenced germination responses in wheat Var-HUW-468 under heavy metal stress

The beneficial effect of seed priming in improving critical growth stages like seed germination and early growth phases has been accepted by Plant Physiologists for many important field crops. In the present investigation, studies were made to see the effect of heavy metal stress imposed during germination using solution of HgCl₂ in four different concentrations (0.0, 0.50, 0.75 and 1.00 mM) in Petri dishes on primed and non-primed seeds of wheat. Priming has been done with distilled water (hydro), Mg(NO₃)₂ and Ca(NO₃)₂ (halo) salts. Different germination parameters, such as germination percentage, radicle and plumule lengths, seedling emergence, soluble and insoluble sugar contents and activity of α-amylase in endosperm were studied at different study periods. Primed seeds increased all the germination parameters except insoluble sugar content in respect to non-primed control in the absence of HgCl₂. However, the use of primed seeds has shown to overcome the inhibitory effects of heavy metal stress imposed in the form of HgCl₂ solution during the period of germination. Hence, the work concludes the mitigating effects of priming under heavy metal stress.