Characterization of different FAD-dependent glucose dehydrogenases for possible use in glucose-based biosensors and biofuel cells

In this study, different flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenases (FADGDHs) were characterized electrochemically after “wiring” them with an osmium redox polymer [Os(4,4′-dimethyl-2,2′-bipyridine)2(PVI)10Cl]+ on graphite electrodes. One tested FADGDH was that recently discovered in Glomerella cingulata (GcGDH), another was the recombinant form expressed in Pichia pastoris (rGcGDH), and the third was a commercially available glycosylated enzyme from Aspergillus sp. (AspGDH). The performance of the Os-polymer “wired” GDHs on graphite electrodes was tested with glucose as the substrate. Optimal operational conditions and analytical characteristics like sensitivity, linear ranges and current density of the different FADGDHs were determined. The performance of all three types of FADGDHs was studied at physiological conditions (pH 7.4). The current densities measured at a 20 mM glucose concentration were 494 ± 17, 370 ± 24, and 389 ± 19 μA cm−2 for GcGDH, rGcGDH, and AspGDH, respectively. The sensitivities towards glucose were 2.16, 1.90, and 1.42 μA mM−1 for GcGDH, rGcGDH, and AspGDH, respectively. Additionally, deglycosylated rGcGDH (dgrGcGDH) was investigated to see whether the reduced glycosylation would have an effect, e.g., a higher current density, which was indeed found. GcGDH/Os-polymer modified electrodes were also used and investigated for their selectivity for a number of different sugars

The Testicular and Epididymal Expression Profile of PLCζ in Mouse and Human Does Not Support Its Role as a Sperm-Borne Oocyte Activating Factor

Phospholipase C zeta (PLCζ) is a candidate sperm-borne oocyte activating factor (SOAF) which has recently received attention as a potential biomarker of human male infertility. However, important SOAF attributes of PLCζ, including its developmental expression in mammalian spermiogenesis, its compartmentalization in sperm head perinuclear theca (PT) and its release into the ooplasm during fertilization have not been established and are addressed in this investigation. Different detergent extractions of sperm and head/tail fractions were compared for the presence of PLCζ by immunoblotting. In both human and mouse, the active isoform of PLCζ was detected in sperm fractions other than PT, where SOAF is expected to reside. Developmentally, PLCζ was incorporated as part of the acrosome during the Golgi phase of human and mouse spermiogenesis while diminishing gradually in the acrosome of elongated spermatids. Immunofluorescence localized PLCζ over the surface of the postacrosomal region of mouse and bull and head region of human spermatozoa leading us to examine its secretion in the epididymis. While previously thought to have strictly a testicular expression, PLCζ was found to be expressed and secreted by the epididymal epithelial cells explaining its presence on the sperm head surface. In vitro fertilization (IVF) revealed that PLCζ is no longer detectable after the acrosome reaction occurs on the surface of the zona pellucida and thus is not incorporated into the oocyte cytoplasm for activation. In summary, we show for the first time that PLCζ is compartmentalized as part of the acrosome early in human and mouse spermiogenesis and is secreted during sperm maturation in the epididymis. Most importantly, no evidence was found that PLCζ is incorporated into the detergent-resistant perinuclear theca fraction where SOAF resides

Regulatory elements and transcriptional control of chicken vasa homologue (CVH) promoter in chicken primordial germ cells

BACKGROUND: Primordial germ cells (PGCs), the precursors of functional gametes, have distinct characteristics and exhibit several unique molecular mechanisms to maintain pluripotency and germness in comparison to somatic cells. They express germ cell-specific RNA binding proteins (RBPs) by modulating tissue-specific cis- and trans-regulatory elements. Studies on gene structures of chicken vasa homologue (CVH), a chicken RNA binding protein, involved in temporal and spatial regulation are thus important not only for understanding the molecular mechanisms that regulate germ cell fate, but also for practical applications of primordial germ cells. However, very limited studies are available on regulatory elements that control germ cell-specific expression in chicken. Therefore, we investigated the intricate regulatory mechanism(s) that governs transcriptional control of CVH. RESULTS: We constructed green fluorescence protein (GFP) or luciferase reporter vectors containing the various 5′ flanking regions of CVH gene. From the 5′ deletion and fragmented assays in chicken PGCs, we have identified a CVH promoter that locates at −316 to +275 base pair fragment with the highest luciferase activity. Additionally, we confirmed for the first time that the 5′ untranslated region (UTR) containing intron 1 is required for promoter activity of the CVH gene in chicken PGCs. Furthermore, using a transcription factor binding prediction, transcriptome analysis and siRNA-mediated knockdown, we have identified that a set of transcription factors play a role in the PGC-specific CVH gene expression. CONCLUSIONS: These results demonstrate that cis-elements and transcription factors localizing in the 5′ flanking region including the 5′ UTR and an intron are important for transcriptional regulation of the CVH gene in chicken PGCs. Finally, this information will contribute to research studies in areas of reproductive biology, constructing of germ cell-specific synthetic promoter for tracing primordial germ cells as well as understanding the transcriptional regulation for maintaining germness in PGCs

Studies on Uranyl Complexes : Part III-Potentiometric Investigation of Uranyl-Amino Acid Complexes

382-38

Toluene methylation activity of zirconium pillared clays

281-285<span style="font-size:11.0pt;line-height:115%; font-family:" calibri","sans-serif";mso-ascii-theme-font:minor-latin;mso-fareast-font-family:="" "times="" new="" roman";mso-fareast-theme-font:minor-fareast;mso-hansi-theme-font:="" minor-latin;mso-bidi-font-family:"times="" roman";mso-ansi-language:en-us;="" mso-fareast-language:en-us;mso-bidi-language:ar-sa"="">Zirconium pillared montmorillonite and vermiculite clay catalysts have been prepared by different methods of intercalation. The catalysts were characterized by XRD, nitrogen adsorption, Infared (IR) and differential scanning calorimetric (DSC) studies. Toluene methylation activity is found to depend on the preparative method and zirconium concentration of the catalysts. The results indicate a direct correlation between the density of Lewis acid sites and toluene methylation activity.</span

Surface characteristics and cracking activity of zirconium pillared clays

119-123Zirconium pillared montmorillonite and vermiculite catalysts have been prepared using four different methods by intercalation with zirconium oxychloride solution. The surface characteristics of the catalysts have been characterized using nitrogen adsorption, XRD and IR studies. The nature of acid site is identified on the basis of IR spectral data of pyridine and ammonia adsorbed zirconium pillared catalysts. The acidity of the samples was determined by butylamine titration as well as by thermal desorption of pyridine and ammonia. Differential scanning calorimetric technique has also been used to probe the nature of acidic sites, their heterogeneity, acid strength and density. Cumene cracking has been carried out as a test reaction of Bronsted acidity. The product selectivity is towards benzene. The cracking activity has been correlated with the density of Bronsted acid sites.</span

Infrared spectral characterization of acidity of zirconium pillared clays

441-443Montmorillonite and vermiculite intercalated with polyoxo cations of zirconium have been prepared. The nature of acidic sites has been studied by infrared (lR) spectroscopy using pyridine and ammonia as the probe molecules. The IR spectral data show three types of acid sites to be available on the pillared clay catalysts. The existence of Bronsted acid, lewis acid and weak acidic sites has been demonstrated. The effect of heat treatment on the intensities of IR spectral bands has been investigated to confirm the presence of weak acid sites to which pyridine is hydrogen bonded

Polarographic Behaviour of Terdentate Hydroxyamino Acid Complexes of Copper(II) Complexes of <img src='http://www.niscair.res.in/jinfo/smaller.gif' border=0><img src='http://www.niscair.res.in/jinfo/small.gif' border=0>-Serine, <img src='http://www.niscair.res.in/jinfo/smaller.gif' border=0><img src='http://www.niscair.res.in/jinfo/small.gif' border=0>-Threonine & <img src='http://www.niscair.res.in/jinfo/smaller.gif' border=0><img src='http://www.niscair.res.in/jinfo/small.gif' border=0>-γ-Amino-β-hydroxybutyric Acid

240-24

Mixed Ligand Complexes of Copper(II) with <img src='http://www.niscair.res.in/jinfo/small.gif' border=0>-Asparagine as Primary Ligand & Imidazole, Histamine & <img src='http://www.niscair.res.in/jinfo/small.gif' border=0>-Histidine as Secondary Ligands

48-5