Reversion mutation of cDNA CA8-204 minigene construct produces a truncated functional peptide that regulates calcium release in vitro and produces profound analgesia in vivo

Mammalian Genome - Intracellular calcium is critical in orchestrating neuronal excitability and analgesia. Carbonic anhydrase-8 (CA8) regulates intracellular calcium signaling through allosteric...     

Systematic review and meta-analysis of PIT tagging effects on mortality and growth of juvenile salmonids

Reviews in Fish Biology and Fisheries - A systematic review and meta-analysis was conducted to evaluate the appropriate tag:fish size ratio when tagging juvenile salmonids (genera Oncorhynchus,...     

Selection of ceramic fluorapatite‐binding peptides from a phage display combinatorial peptide library: optimum affinity tags for fluorapatite chromatography

Peptide affinity tags have become efficient tools for the purification of recombinant proteins from biological mixtures. The most commonly used ligands in this type of affinity chromatography are immobilized metal ions, proteins, antibodies, and complementary peptides. However, the major bottlenecks of this technique are still related to the ligands, including their low stability, difficulties in immobilization, and leakage into the final products. A model approach is presented here to overcome these bottlenecks by utilizing macroporous ceramic fluorapatite (CFA) as the stationary phase in chromatography and the CFA‐specific short peptides as tags. The CFA chromatographic materials act as both the support matrix and the ligand. Peptides that bind with affinity to CFA were identified from a randomized phage display heptapeptide library. A total of five rounds of phage selection were performed. A common N‐terminal sequence was found in two selected peptides: F4‐2 (KPRSMLH) and F5‐4 (KPRSVSG). The peptide F5‐4, displayed by more than 40% of the phages analyzed in the fifth round of selection, was subjected to further studies. Selectivity of the peptide for the chemical composition and morphology of CFA was assured by the adsorption studies. The dissociation constant, obtained from the F5‐4/CFA adsorption isotherm, was in the micromolar range, and the maximum capacity was 39.4 nmol/mg. The chromatographic behavior of the peptides was characterized on a CFA stationary phase with different buffers. Preferential affinity and specific retention properties suggest the possible application of the phage‐derived peptides as a tag in CFA affinity chromatography for enhancing the selective recovery of proteins. Copyright © 2013 John Wiley & Sons, Ltd.     

Comparative analysis of the methods used for finding surface energy to investigate protein interaction behavior on chromatographic supports

Hydrophobic interaction chromatography, an important and effective purification strategy, is generally used for the purification of variety of biomolecules. A basic understanding of the protein interaction behavior is required to effectively separate these biomolecules. A colloidal type extended Derjaguin, Landau, Verwey, and Overbeek calculations were utilized to study the interactions behavior of model proteins to commercially available hydrophobic chromatographic materials that is, Toyopearl Phenyl 650C and Toyopearl Butyl 650C. Physicochemical properties of selected model proteins were achieved by contact angle and zeta potential measurements. The contact angle of chromatographic materials used was achieved through sessile drop method on disrupted beads and capillary penetration method (CPM) on intact beads. The surface properties were further used to calculate the interactions of the proteins to chromatographic supports. The calculated secondary energy minimum of the proteins with the chromatographic materials (from the contact angle values determined through both methods can be correlated with the retention volumes from the real chromatography. The secondary energy minimum values are higher for each protein to the chromatographic materials calculated from the inputs derived through sessile drop method compared to CPM. For instance, immunoglobulin G has secondary energy minimum value of 0.17 kT compared to 0.11 kT, obtained through sessile drop method and CPM, respectively. Average relative values of the energy minimum calculated for all proteins are as 1.51 kT and 1.29 kT for Toyopearl Butyl 650C and Toyopearl Phenyl 650C, respectively, as a conversion factor for estimation of secondary energy minimum for both methods.     

Synthesis and performance of 3D-megaporous structures for enzyme immobilization and protein capture

The preparation of megaporous bodies, with potential applications in biotechnology, was attempted by following several strategies. As a first step, naive and robust scaffolds were produced by polymerization of selected monomers in the presence of a highly soluble cross‐linker agent. Ion‐exchange function was incorporated by particle embedding, direct chemical synthesis, or radiation‐induced grafting. The total ionic capacity of such systems was 1.5 mmol H⁺/g, 1.4 mmol H⁺/g, and 17 mmol H⁺/g, respectively. These values were in agreement with the ability to bind model proteins: observed dynamic binding capacity at 50% breakthrough was ?7.2 mg bovine serum albumin/g, ?7.4 hen egg‐white lysozyme (HEWL) mg/g, and ?108 HEWL mg/g. In the later case, total (static) binding capacity reached 220 mg/g. It was observed that the structure and size of the megapores remained unaffected by the grafting procedure which, however, allowed for the highest protein binding capacity. Lysozyme supported on grafted body showed extensive clarification activity against a Micrococcus lysodekticus suspension in the flow‐through mode, i.e., 90% destruction of suspended microbial cells was obtained with a residence time ≈ 18 min. Both protein capture and biocatalysis applications are conceivable with the 3D‐megaporous materials described in this work. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

Metal-based new sulfonamides: Design, synthesis, antibacterial, antifungal, and cytotoxic properties

Cobalt(II), copper(II), nickel(II) and zinc(II) metal complexes with 5-chlorosalicyladehyde derived Schiff base sulfonamides have been synthesized and characterized. Structure and bonding nature of all the synthesized compounds have been deduced from physical, analytical, and spectral (IR, (1)H NMR, (13)C NMR, Mass, electronic) data. An octahedral geometry has been proposed for all the metal complexes. The ligands and their metal complexes have been screened for their in vitro antibacterial, antifungal, and cytotoxic properties and results are reported.     

Synthesis, characterization and biological studies of sulfonamide Schiff's bases and some of their metal derivatives

A new series of Schiff base ligands derived from sulfonamide and their metal(II) complexes [cobalt(II), copper(II), nickel(II) and zinc(II)] have been synthesized and characterized. The nature of bonding and structure of all the synthesized compounds has been explored by physical, analytical and spectral data of the ligands and their metal(II) complexes. The authors suggest that all the prepared complexes possess an octahedral geometry. The ligands and metal(II) complexes have been screened for their in vitro antibacterial activity against bacterial strains, Escherichia coli, Shigella flexneri, Pseudomonas aeruginosa, Salmonella typhi and for antifungal activity against fungal strains, Trichophyton longifusus, Candida albicans, Aspergillus flavus, Microsporum canis, Fusarium solani and Candida glabrata. These assays enabled the identification of the metal complexes as an effective antimicrobial agent with low cytotoxicity.     

Genetically determined P2X7 receptor pore formation regulates variability in chronic pain sensitivity

Chronic pain is highly variable between individuals, as is the response to analgesics. Although much of the variability in chronic pain and analgesic response is heritable, an understanding of the genetic determinants underlying this variability is rudimentary. Here we show that variation within the coding sequence of the gene encoding the P2X7 receptor (P2X7R) affects chronic pain sensitivity in both mice and humans. P2X7Rs, which are members of the family of ionotropic ATP-gated receptors, have two distinct modes of function: they can function through their intrinsic cationic channel or by forming nonselective pores that are permeable to molecules with a mass of up to 900 Da. Using genome-wide linkage analyses, we discovered an association between nerve-injury-induced pain behavior (mechanical allodynia) and the P451L mutation of the mouse P2rx7 gene, such that mice in which P2X7Rs have impaired pore formation as a result of this mutation showed less allodynia than mice with the pore-forming P2rx7 allele. Administration of a peptide corresponding to the P2X7R C-terminal domain, which blocked pore formation but not cation channel activity, selectively reduced nerve injury and inflammatory allodynia only in mice with the pore-forming P2rx7 allele. Moreover, in two independent human chronic pain cohorts, a cohort with pain after mastectomy and a cohort with osteoarthritis, we observed a genetic association between lower pain intensity and the hypofunctional His270 (rs7958311) allele of P2RX7. Our findings suggest that selectively targeting P2X7R pore formation may be a new strategy for individualizing the treatment of chronic pain.     

Determination of the Genetic and Synergistic Suppression of a Methoxyfenozide-Resistant Strain of the House Fly <Emphasis Type="Italic">Musca domestica</Emphasis> L. (Diptera: Muscidae)

Musca domestica Linnaeus (house fly, Diptera: Muscidae) is a major veterinary and medical important pest all over the world. These flies have ability to develop resistance to insecticides. The present trial was performed to discover the inheritance mode (autosomal, dominance, number of genes involved) and preliminary mechanism of methoxyfenozide resistance in order to provide basic information necessary to develop resistance management strategy for this pest. A strain of M. domestica (MXY-SEL) was exposed to methoxyfenozide for 44 generations which developed a 5253.90-fold level of resistance to methoxyfenozide. The overlapping fiducial limits of LC₅₀ values of the reciprocal crosses, F₁ (MXY-SEL ♂?×?Susceptible ♀) and F₁^? (MXY-SEL ♀?×?Susceptible ♂), suggest that inheritance of methoxyfenozide resistance was an autosomal and likely completely dominant trait (D_LC?=?0.93 and 0.94 for F₁ and F₁^?, respectively). Backcrosses of the F₁ with the parental MXY-SEL or Susceptible population predict a polygenic mode of inheritance. Piperonyl butoxide significantly altered the LC₅₀ values, suggesting enhanced detoxification by cytochrome P450-dependent monooxygenases is a major mechanism of resistance to methoxyfenozide in the MXY-SEL strain. The estimated realized heritability was 0.07 for methoxyfenozide. These results would be helpful for the better management of M. domestica.