Enzyme-linked immunosorbent assay of changes in serum levels of growth hormone (cGH) in common carps (Cyprinus carpio)

The aim of the present study was to purify the common native carp growth hormone (ncGH), produce monoclonal antibodies (mAbs) to common native carp growth hormone (ncGH), and further enhance the sensitivity of enzyme-linked immunosorbent assays (ELISA) for ncGH. Additionally, we investigated changes in serum ncGH levels in carps raised in different environmental conditions. The recombinant grass carp (Ctenopharyngodon idella) growth hormone was purified and used as antigen to immunize the rabbit. The natural ncGH was isolated from the pituitaries of common carp. SDS-PAGE and Western blot utilizing the polyclonal anti-rgcGH antibody confirmed the purification of ncGH from pituitaries. Purified ncGH was then used as an immunogen in the B lymphocyte hybridoma technique. A total of 14 hybridoma cell lines (FMU-cGH 1-14) were established that were able to stably secrete mAbs against ncGH. Among them, eight clones (FMU-cGH1-6, 12 and 13) were successfully used for Western blot while nine clones (FMU-cGH 1-7, 9 and 10) were used in fluorescent staining and immunohistochemistry. Epitope mapping by competitive ELISA demonstrated that these mAbs recognized five different epitopes. A sensitive sandwich ELISA for detection of ncGH was developed using FMU-cGH12 as the coating mAb and FMU-cGH6 as the enzyme labeled mAb. This detection system was found to be highly stable and sensitive, with detection levels of 70 pg/mL. Additionally, we found that serum ncGH levels in restricted food group and in the net cage group increased 6.9-and 5.8-fold, respectively, when compared to controls, demonstrating differences in the GH stress response in common carp under different living conditions.The aim of the present study was to purify the common native carp growth hormone (ncGH), produce monoclonal antibodies (mAbs) to common native carp growth hormone (ncGH), and further enhance the sensitivity of enzyme-linked immunosorbent assays (ELISA) for ncGH. Additionally, we investigated changes in serum ncGH levels in carps raised in different environmental conditions. The recombinant grass carp (Ctenopharyngodon idella) growth hormone was purified and used as antigen to immunize the rabbit. The natural ncGH was isolated from the pituitaries of common carp. SDS-PAGE and Western blot utilizing the polyclonal anti-rgcGH antibody confirmed the purification of ncGH from pituitaries. Purified ncGH was then used as an immunogen in the B lymphocyte hybridoma technique. A total of 14 hybridoma cell lines (FMU-cGH 1-14) were established that were able to stably secrete mAbs against ncGH. Among them, eight clones (FMU-cGH1-6, 12 and 13) were successfully used for Western blot while nine clones (FMU-cGH 1-7, 9 and 10) were used in fluorescent staining and immunohistochemistry. Epitope mapping by competitive ELISA demonstrated that these mAbs recognized five different epitopes. A sensitive sandwich ELISA for detection of ncGH was developed using FMU-cGH12 as the coating mAb and FMU-cGH6 as the enzyme labeled mAb. This detection system was found to be highly stable and sensitive, with detection levels of 70 pg/mL. Additionally, we found that serum ncGH levels in restricted food group and in the net cage group increased 6.9-and 5.8-fold, respectively, when compared to controls, demonstrating differences in the GH stress response in common carp under different living conditions

Amide N-glycosylation by Asm25, an N-glycosyltransferase of ansamitocins

Ansamitocins are potent antitumor maytansinoids produced by Actinosynnema pretiosum. Their biosynthesis involves the initial assembly of a macrolactam polyketide, followed by a series of postpolyketide synthase (PKS) modifications. Three ansamitocin glycosides were isolated from A. pretiosum and fully characterized structurally as novel ansamitocin derivatives, carrying a (3-D-glucosyl group attached to the macrolactam amide nitrogen in place of the N-methyl group. By gene inactivation and complementation, asm25 was identified as the N-glycosyltransferase gene responsible for the macrolactam amide N-glycosylation of ansamitocins. Soluble, enzymatically active Asm25 protein was obtained from asm25-expressing E, coli by solubilization from inclusion bodies. Its optimal reaction conditions, including temperature, pH, metal ion requirement, and Km/Kcat, were determined. Asm25 also showed broad substrate specificity toward other ansamycins and synthetic indolin-2-ones. To the best of our knowledge, this represents the first in vitro characterization of a purified antibiotic N-glycosyltransferase.National Natural Science Fund for Distinguished Young Scholars [30325044]; Natural Science Foundation of China [30600005, 30570019]; Ministry of Science and Technolog

Ethanol electrooxidation on carbon supported PtSn catalyst: In situ TRFTIR study

Carbon supported PtSn catalyst (PtSn/C) was prepared by a modified polyol method and characterized by means of XRD. It was showed that the metal particle size was 2.2 nm and the unit cell parameter increased compared with Pt/C. In situ time-resolved Fourier transform infrared spectroscopy (TRFTIRS) was used to study the electrooxidation of ethanol on PtSn/C catalyst. CO was the main poison species adsorbed on the active sites to inhibit the further reaction of ethanol electrooxidation. Acetaldehyde and acetic acid were found to be the products of ethanol electrooxidation as competing reactions with ethanol dissociation when the potential was up to 0.3 V, which reduced the poisoning effect. The selectivity of acetic acid among the products was improved with the increase in the potential and reaction time. CO2, which appeared at 0.4 V I was the final product and yielded from the oxidation of COL. The catalytic mechanism of PtSn/C towards ethanol electrooxidation was analyzed based on the results

Phylogeny of Chinese catfishes inferred from mitochondrial cytochrome b sequences

The mitochondrial DNA cytochrome b gene was sequenced from 27 catfish species representing 11 families and 24 genera catfishes in China. Aligning with cytochrome b sequences of eight catfish species from North America and Africa retrieved from GenBank,and selecting Astyanax mexicanu, Cyprinus carpio,and Sardinops melanostictus as outgroups, we constructed a matrix of 38 DNA sequences. The phylogenetic trees were constructed by using Bayesian method and Maximum Parsimony ( MP) method. The results showed that ( 1) there are three base pair deletions of mitochondrial cytochrome b gene compared with Characiformes,Cypriniformes-and Clupeiformes; (2) the representatives of Chinese catfish species form a monophyletic group; and (3) the molecular phylogenetic trees constructed with both methods suggest that the families Sisoridae, Akysidae and Amblycipitidae form a monophyletic group, and the families Clariidae, Schilbidae, Ariidae, Ictaluridae, Cranoglanididae, Pangasiidae, Siluridae, Claroteidae, and Bagridae also form a monophyletic group. The families Cranoglanididae from China and Ictaluridae from North America form a sister-group relationship, and the families Clariidae. Ictaluridae, Siluridae, and Sisoridae are obviously monophyletic groups. But the position of the family Plotosidae was not resolved by Bayesian analysis and maximum parsimony inference

Theoretical Studies on Thermochemistry for Conversion of 5-Chloromethylfurfural into Valuable Chemicals

Recently, 5-chloromethylfurfural (CMF) was proposed as a central, intermediate in the conversion of carbohydrate-based material into useful organic commodities. In the present work, we have calculated the thermochemistry using the highly accurate G4 theory and several state-of-art density functional theory (DFT) methods (e.g., x1, M06-2X, B2PLYP-D, and XYG3) for the conversion from CMF to 5-hydroxymethylfurfural (HMF) and levulinic acid (LA) in water, and that to biofuels 5-ethoxymethylfurfural (EMF) and ethyllevulinate (EL) in alcohol. New reaction mechanisms have been proposed, which complement the well-recognized Horvat mechanisms. The assessment of DFT methods suggested that XYG3 be a viable method for biomass related thermochemistry calculations.National Natural Science Foundation of China[91027044, 20973138, 21133004]; Ministry of Science and Technology of China[2007CB815206, 2011CB808505]; Synfuels China Co. Ltd

Design of two-dimensional photonic crystal edge emitting laser for photonic integrated circuits

An edge emitting laser based on two-dimensional photonic crystal slabs is proposed. The device consists of a square lattice microcavity, which is composed of two structures with the same period but different radius of air-holes, and a waveguide. In the cavity, laser resonance in the inner structure benelits from not only the anomalous dispersion characteristic of the first band-edge at the M point in the first Brillouin-zone but also zero photon states in the outer structure. A line defect waveguide is introduced in the outer structure for extracting photons from the inner cavity. Three-dimensional finite-difference time-domain simulations apparently show the in-plane laser output from the waveguide. The microcavity has an effective mode volume of about 3.2(lambda/eta(slab))(3) for oscillation -mode and the quality factor of the device including line defect waveguide is estimated to be as high as 1300

Sept6 Is Required for Ciliogenesis in Kupffer's Vesicle, the Pronephros, and the Neural Tube during Early Embryonic Development

Septins are conserved filament-forming GTP-binding proteins that act as cellular scaffolds or diffusion barriers in a number of cellular processes. However, the role of septins in vertebrate development remains relatively obscure. Here, we show that zebrafish septin 6 (sept6) is first expressed in the notochord and then in nearly all of the ciliary organs, including Kupffer's vesicle (KV), the pronephros, eye, olfactory bulb, and neural tube. Knockdown of sept6 in zebrafish embryos results in reduced numbers and length of cilia in KV. Consequently, cilium-related functions, such as the left-right patterning of internal organs and nodal/spaw signaling, are compromised. Knockdown of sept6 also results in aberrant cilium formation in the pronephros and neural tube, leading to cilium-related defects in pronephros development and Sonic hedgehog (Shh) signaling. We further demonstrate that SEPT6 associates with acetylated alpha-tubulin in vivo and localizes along the axoneme in the cilia of zebrafish pro-nephric duct cells as well as cultured ZF4 cells. Our study reveals a novel role of sept6 in ciliogenesis during early embryonic development in zebrafish.Septins are conserved filament-forming GTP-binding proteins that act as cellular scaffolds or diffusion barriers in a number of cellular processes. However, the role of septins in vertebrate development remains relatively obscure. Here, we show that zebrafish septin 6 (sept6) is first expressed in the notochord and then in nearly all of the ciliary organs, including Kupffer's vesicle (KV), the pronephros, eye, olfactory bulb, and neural tube. Knockdown of sept6 in zebrafish embryos results in reduced numbers and length of cilia in KV. Consequently, cilium-related functions, such as the left-right patterning of internal organs and nodal/spaw signaling, are compromised. Knockdown of sept6 also results in aberrant cilium formation in the pronephros and neural tube, leading to cilium-related defects in pronephros development and Sonic hedgehog (Shh) signaling. We further demonstrate that SEPT6 associates with acetylated alpha-tubulin in vivo and localizes along the axoneme in the cilia of zebrafish pro-nephric duct cells as well as cultured ZF4 cells. Our study reveals a novel role of sept6 in ciliogenesis during early embryonic development in zebrafish

Synthesis and antioxidant ability of 6,6 '-diamino-6,6 '-dideoxytrehalose

In order to study the influence of amino group on antioxidant activity of oligosaccharides, an amino disaccharide, 6,6 '-diamino-6,6 '-dideoxytrehalose (DAMDT) was successfully prepared in this paper, and its antioxidant activities against DPPH, superoxide, hydrogen peroxide, and hydroxyl radicals, and reducing power were evaluated, respectively. The results indicated that DAMDT had better antioxidant activity than trehalose at any tested concentration. The influence of amino group on antioxidant activity of disaccharides is positive based on the results in this paper, and amination should be an effective method to improve the bioactivity of saccharides. (C) 2017 Elsevier Inc. All rights reserved

Antioxidant activity of inulin derivatives with quaternary ammonium

Three inulin derivatives with quaternary ammonium, including 2-(N, N, N-triethylamine)acetyl inulin chloride (TAIL), 2-imidazoleacetyl inulin chloride (IAIL), and 2-(1-methylimidazole)acetyl inulin chloride (MIAIL) were synthesized via reaction of chloracetyl inulin (CAIL) with tertiary amine. Their antioxidant activity against hydroxyl radicals and superoxide radicals were evaluated in vitro, respectively. The results showed that IAIL had the best scavenging ability on scavenging hydroxyl radicals and superoxide radicals, and the scavenging indices were 86.7 and 67.8% at 1.6 mg/mL, respectively. These data indicate that all of the inulin derivatives have better antioxidant activities than inulin, and the quaternary ammonium and imidazole will improve the antioxidant activity of inulin derivatives

Repression of hypoxia-inducible factor alpha signaling by Set7-mediated methylation

Hypoxia-inducible factor (HIF)-1 alpha and HIF-2 alpha are the main regulators of cellular responses to hypoxia. Post-translational modifications of HIF-1 alpha and 2 alpha are necessary to modulate their functions. The methylation of non-histone proteins by Set7, an SET domain-containing lysine methyltransferase, is a novel regulatory mechanism to control cell protein function in response to various cellular stresses. In this study, we show that Set7 methylates HIF-1 alpha at lysine 32 and HIF-2 alpha at lysine K29; this methylation inhibits the expression of HIF-1 alpha/2 alpha targets by impairing the occupancy of HIF-alpha on hypoxia response element of HIF target gene promoter. Set7-null fibroblasts and the cells with shRNA-knocked down Set7 exhibit upregulated HIF target genes. Set7 inhibitor blocks HIF-1 alpha/2 alpha methylation to enhance HIF target gene expression. Set7-null fibroblasts and the cells with shRNA-knocked down Set7 or inhibition of Set7 by the inhibitor subjected to hypoxia display an increased glucose uptake and intracellular adenosine triphosphate levels. These findings define a novel modification of HIF-1 alpha/2 alpha and demonstrate that Set7-medited lysine methylation negatively regulates HIF-alpha transcriptional activity and HIF-1 alpha-mediated glucose homeostasis