Sample records for melibiose

  1. Efficiency Analysis and Mechanism Insight of that Whole-Cell Biocatalytic Production of Melibiose from Raffinose with Saccharomyces cerevisiae. (United States)

    Zhou, Yingbiao; Zhu, Yueming; Dai, Longhai; Men, Yan; Wu, Jinhai; Zhang, Juankun; Sun, Yuanxia


    Melibiose is widely used as a functional carbohydrate. Whole-cell biocatalytic production of melibiose from raffinose could reduce its cost. However, characteristics of strains for whole-cell biocatalysis and mechanism of such process are unclear. We compared three different Saccharomyces cerevisiae strains (liquor, wine, and baker's yeasts) in terms of concentration variations of substrate (raffinose), target product (melibiose), and by-products (fructose and galactose) in whole-cell biocatalysis process. Distinct difference was observed in whole-cell catalytic efficiency among three strains. Furthermore, activities of key enzymes (invertase, α-galactosidase, and fructose transporter) involved in process and expression levels of their coding genes (suc2, mel1, and fsy1) were investigated. Conservation of key genes in S. cerevisiae strains was also evaluated. Results show that whole-cell catalytic efficiency of S. cerevisiae in the raffinose substrate was closely related to activity of key enzymes and expression of their coding genes. Finally, we summarized characteristics of producing strain that offered advantages, as well as contributions of key genes to excellent strains. Furthermore, we presented a dynamic mechanism model to achieve some mechanism insight for this whole-cell biocatalytic process. This pioneering study should contribute to improvement of whole-cell biocatalytic production of melibiose from raffinose.

  2. Construction of engineered Saccharomyces cerevisiae strain to improve that whole-cell biocatalytic production of melibiose from raffinose. (United States)

    Zhou, Yingbiao; Zhu, Yueming; Men, Yan; Dong, Caixia; Sun, Yuanxia; Zhang, Juankun


    There are excessive by-products in the biocatalysis process of this whole-cell biocatalytic production of melibiose from raffinose with current Saccharomyces cerevisiae strains. To solve this problem, we constructed engineered strains based on a liquor yeast (S. cerevisiae) via gene deletion (mel1 gene), heterologous integration (fsy1 or/and ffzi1 gene from Candida magnoliae), and gene overexpression (gcr1 gene). Functional verification showed that deletion of the mel1 gene led to elimination of the reactions catalyzed by α-galactosidase, as well as elimination of the degradation of melibiose and the formation of galactose by-product. Insertion of the fsy1 or/and ffzi1 gene and overexpression of the gcr1 gene could contribute to fructose transport for enhancing the biopurification rate of the fructose by-product. Compared with the wild-type strain, the optimal engineered strain of MP8 (Δmel1::fsy1 cm ::ffzi1 cm ::gcr1 sc ) had improved about 30% on yield, 31% on productivity, and 36% on purity of the melibiose product.

  3. The potential of lactulose and melibiose, two novel trehalase-indigestible and autophagy-inducing disaccharides, for polyQ-mediated neurodegenerative disease treatment. (United States)

    Lee, Guan-Chiun; Lin, Chih-Hsin; Tao, Yu-Chen; Yang, Jinn-Moon; Hsu, Kai-Cheng; Huang, Yin-Jung; Huang, Shih-Han; Kung, Pin-Jui; Chen, Wan-Ling; Wang, Chien-Ming; Wu, Yih-Ru; Chen, Chiung-Mei; Lin, Jung-Yaw; Hsieh-Li, Hsiu Mei; Lee-Chen, Guey-Jen


    The unique property of trehalose encourages its pharmaceutical application in aggregation-mediated neurodegenerative disorders, including Alzheimer's, Parkinson's, and many polyglutamine (polyQ)-mediated diseases. However, trehalose is digested into glucose by trehalase and which reduced its efficacy in the disease target tissues. Therefore, searching trehalase-indigestible analogs of trehalose is a potential strategy to enhance therapeutic effect. In this study, two trehalase-indigestible trehalose analogs, lactulose and melibiose, were selected through compound topology and functional group analyses. Hydrogen-bonding network analyses suggest that the elimination of the hydrogen bond between the linker ether and aspartate 321 (D321) of human trehalase is the key for lactulose and melibiose to avoid the hydrolyzation. Using polyQ-mediated spinocerebellar ataxia type 17 (SCA17) cell and slice cultures, we found the aggregation was significantly prohibited by trehalose, lactulose, and melibiose, which may through up-regulating of autophagy. These findings suggest the therapeutic applications of trehalase-indigestible trehalose analogs in aggregation-associated neurodegenerative diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Structural studies of the melibiose permease of Escherichia coli by fluorescence resonance energy transfer. I. Evidence for ion-induced conformational change. (United States)

    Maehrel, C; Cordat, E; Mus-Veteau, I; Leblanc, G


    Further insight into the cosubstrate-induced structural change of the melibiose permease (MelB) of Escherichia coli has been sought by investigating the binding and spectroscopic properties of the fluorescent sugar 2'-(N-5-dimethylaminonaphthalene-1-sulfonyl)aminoethyl 1-thio-beta-D-galactopyranoside (Dns2-S-Gal) and related analogs (Dns3-S-Gal or Dns6-S-Gal with a propyl or hexyl instead of an ethyl linker, respectively) interacting with MelB in membrane vesicles or in proteoliposomes. The three analogs efficiently inhibit melibiose transport and bind to MelB in a sodium-dependent fashion. Their dissociation constants (Kd) are in the micromolar range in the presence of NaCl and an order of magnitude higher in its absence. In the presence of NaCl and Dns2-S-Gal, sample excitation at 335 or 297 nm gives rise to a fluorescent signal at around 465 nm, whereas Dns3-S-Gal or Dns6-S-Gal emits a fluorescence light at 490 or 506 nm, respectively. Detailed study of the Dns2-S-Gal signal elicited by a 297 nm illumination indicates that a tryptophan-mediated fluorescence resonance energy transfer phenomenon is involved in the response. All fluorescence signals below 500 nm are prevented by addition of melibiose in excess, and the kinetic constants describing their dependence on the probe or NaCl concentrations closely correlate with the probe binding constants. Finally, the Dns2-S-Gal signal recorded in sodium-free medium is red shifted by up to 25 nm from that recorded in the presence of NaCl. Taken together, these results suggest (i) that the fluorescence signals below 500 nm arise from Dns-S-Gal molecules bound to MelB, (ii) the presence of a highly hydrophobic environment close to or at the sugar-binding site, the polarity of which increases on moving away from the sugar-binding site, and (iii) that the interaction of sodium ions with MelB enhances the hydrophobicity of this environment. These results are consistent with the induction of a cooperative change of the

  5. Lactose Transport System of Streptococcus thermophilus : a Hybrid Protein with Homology to the Melibiose Carrier and Enzyme III of Phosphoenolpyruvate-Dependent Phosphotransferase Systems

    NARCIS (Netherlands)

    Poolman, Bert; Royer, Theresa J.; Mainzer, Stanley E.; Schmidt, Brian F.


    The gene responsible for the transport of lactose into Streptococcus thermophilus (lacS) was cloned in Escherichia coli as a 4.2-kilobase fragment from an EcoRI library of chromosomal DNA by using the vector pKK223-3. From deletion analysis, the gene for lactose transport mapped to two HindIII

  6. Gustatory perception and metabolic utilization of sugars by Myrmica rubra ant workers

    NARCIS (Netherlands)

    Boevé, J-L.; Wäckers, F.L.


    The suitability of various nectar and honeydew sugars as a food source for the polyphagous ant species M. rubra (L.) was studied. The sugars used included monosaccharides (fructose, glucose, galactose, mannose, rhamnose), disaccharides (sucrose, maltose, trehalose, melibiose, lactose) and

  7. Increasing the transglycosylation activity of alpha-galactosidase from Bifidobacterium adolescentis DSM 20083 by site-directed mutagenesis

    NARCIS (Netherlands)

    Hinz, S.W.A.; Doeswijk-Voragen, C.H.L.; Schipperus, R.; Broek, van den L.A.M.; Vincken, J.P.; Voragen, A.G.J.


    The ¿-galactosidase (AGA) from Bifidobacterium adolescentis DSM 20083 has a high transglycosylation activity. The optimal conditions for this activity are pH 8, and 37°C. At high melibiose concentration (600 mM), approximately 64% of the enzyme-substrate encounters resulted in transglycosylation.

  8. Lectins, Mitogenicity and Seed Germination: A Comparative Study ...

    African Journals Online (AJOL)

    Dr J. T. Ekanem


    May 24, 2007 ... generally, e. g. Lactose, but also by Galactose. Secondly, the seeds of A. communis, a plant belonging to the family Moraceae contain a mitogenic haemagglutinin (Artocarpin) that is specifically inhibited by the Melibiose, (6-O-α-. D-galactopyranosyl-D-glucose), but neither by. Lactose nor Galactose4.

  9. Microbial Biosensors for Selective Detection of Disaccharides (United States)

    Seven microbial strains were screened for their ability to detect disaccharides as components of Clark-type oxygen biosensors. Sensors responded to varying degrees to maltose, cellobiose, sucrose, and melibiose, but none responded strongly to lactose. Although microbial sensors are relatively nons...

  10. Purification and characterization of an alpha-galactosidase from Aspergillus fumigatus

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    Sebastião Tavares de Rezende


    Full Text Available Aspergillus fumigatus secreted invertase (beta-fructofuranosidase and alpha-galactosidase enzymatic activities able to hydrolyzing raffinose oligosaccharides (RO. alpha-Galactosidase was induced by galactose, melibiose and raffinose, but galactose was the most efficient inducer. It was purified by gel filtration and two ion exchange chromatographies and showed Mw of 54.7 kDa. The purified enzyme showed maximal activity against p-nitrophenyl-alpha-D-galactopyranoside (pNPGal at pH 4.5-5.5 and 55 °C, and retained about 80% of the original activity after incubation for 90 minutes at 50ºC. The KM for pNPGal was 0.3 mM. Melibiose was hydrolyzed by the enzyme but raffinose was very poor substrate.O fungo termofílico Aspergillus fumigatus secreta as enzimas invertase (beta-frutofuranosidase e alfa-galactosidase (alfa-D-galactosídeo galactohi-drolase que estão envolvidas na hidrólise completa dos oligossacarídeos de rafinose. A enzima alfa-galactosidase foi produzida em meio de cultura do fungo Aspergillus fumigatus crescido por 36 h a 42 °C em meio mineral mínimo contendo os açúcares galactose, ou melibiose, ou rafinose como fontes de carbono. A enzima foi purificada por filtração em gel, seguida por duas cromatografias de troca iônica. A massa molecular da alfa-galactosidase determinada por SDS-PAGE foi de 54,7 kDa. A atividade máxima da enzima purificada, utilizando o substrato ro-nitrofenil-alfa-D-galactopiranosídeo (roNPGal foi na faixa de pH entre 4,5 e 5,5 e a 55 °C. A enzima manteve aproximadamente 80% de sua atividade original mesmo após pré-incubação por 90 minutos a 50 °C. O valor de KM para o substrato roNPGal foi 0,3 mM. A enzima foi capaz de hidrolisar melibiose, mas sua atividade foi muito reduzida na presença do substrato rafinose.

  11. Caractérisation physico-chimique et sensorielle de diverses ...

    African Journals Online (AJOL)

    Glucosamine; le Cis-Inositol; le D-Threitol et le Melibiose. La Figure 5 rend compte de la composition qualitative des sucres de la formulation F4 préférée par les dégustateurs. Plusieurs oses et dérivés ont été identifiés parmi lesquels: le Methyl 3-o-acetyl-2,4-di-o-methyl-α-. D-Xylopyranoside ; le D-Glucosamine ; le.

  12. [Effect of lactic acid bacteria isolated from Tibetan Plateau on silage fermentation quality of Elms nutans]. (United States)

    Zhang, Hongmei; Ke, Wencan; Jing, Peixin; Zhang, Juan; Chen, Ming; Yu, Yingwen; Guo, Xusheng


    In order to detect the effect of lactic acid bacteria isolated from Tibetan Plateau on silage fermentation quality of Elms nutans. We used 3 isolated lactic acid bacteria with better growth at low temperatures of 10 and 15 degrees C at ensiling of Elymus nutans. Subsequently, effects of the selected lactic acid bacteria on fermentation profiles of Elymus nutans silages stored at 15 and 25 degrees C were evaluated by using the same species of commercial inoculants as the control. PP-6 isolated from Tibetan Plateau could ferment raffinose, lactose, sorbitol, melibiose and sucrose, and LS-5 could ferment cottonseed sugar, laetrile, rhamnose, lactose, sorbitol, xylose, arabinose, melibiose and sucrose, but the same species of commercial strains could not use these sugars. Inoculation of these three strains into Elymus nutans at 15 and 25 degrees C ensiled for 50 d, we found that LS-5 significantly reduced silage pH, propionic acid concentration and ratio of ammonia nitrogen/total nitrogen at 15 degrees C (P lactic acid concentration (P < 0.05), resulting in a lower ratio of ammonia nitrogen/total nitrogen, saved more crude protein and significantly reduced neutral detergent fiber content (P < 0.05) as compared with the commercial strains. The three isolated strains can improve silage quality of Elymus nutans growing on the Qinghai-Tibetan Plateau at low temperature, but these strains have no obvious advantages at 25 degrees C in comparison with the commercial inoculants.

  13. Truncation of Gal4p explains the inactivation of the GAL/MEL regulon in both Saccharomyces bayanus and some Saccharomyces cerevisiae wine strains. (United States)

    Dulermo, Rémi; Legras, Jean-Luc; Brunel, François; Devillers, Hugo; Sarilar, Véronique; Neuvéglise, Cécile; Nguyen, Huu-Vang


    In the past, the galactose-negative (Gal(-)) phenotype was a key physiological character used to distinguish Saccharomyces bayanus from S. cerevisiae In this work, we investigated the inactivation of GAL gene networks in S. bayanus, which is an S. uvarum/S. eubayanus hybrid, and in S. cerevisiae wine strains erroneously labelled 'S. bayanus'. We made an inventory of their GAL genes using genomes that were either available publicly, re-sequenced by us, or assembled from public data and completed with targeted sequencing. In the S. eubayanus/S. uvarum CBS 380(T) hybrid, the GAL/MEL network is composed of genes from both parents: from S. uvarum, an otherwise complete set that lacks GAL4, and from S. eubayanus, a truncated version of GAL4 and an additional copy of GAL3 and GAL80 Similarly, two different truncated GAL4 alleles were found in S. cerevisiae wine strains EC1118 and LalvinQA23. The lack of GAL4 activity in these strains was corrected by introducing a full-length copy of S. cerevisiae GAL4 on a CEN4/ARS plasmid. Transformation with this plasmid restored galactose utilisation in Gal(-) strains, and melibiose fermentation in strain CBS 380(T) The melibiose fermentation phenotype, formerly regarded as characteristic of S. uvarum, turned out to be widespread among Saccharomyces species. © FEMS 2016. All rights reserved. For permissions, please e-mail:

  14. Identification and quantitative analysis of stage-specific carbohydrates in loblolly pine (Pinus taeda) zygotic embryo and female gametophyte tissues. (United States)

    Pullman, Gerald S; Buchanan, Mike


    Stage-specific analyses of starch and 18 sugars, including pentoses, hexoses, disaccharides, trisaccharides, oligosaccharides and sugar alcohols, were made throughout seed development for zygotic embryo and female gametophyte (FG) tissues of loblolly pine (Pinus taeda L.). Tissue was most often analyzed in triplicate from two open-pollinated families grown in different locations and sampled in different years. Carbohydrates were analyzed by enzymatic assay, high performance liquid chromatography or gas chromatography/mass spectrometry. For all carbohydrates quantified, peak concentrations were higher in embryo tissue than in FG tissue. Significant changes in starch and sugar concentrations occurred over time, with both seed collections showing similar trends in temporal changes. Although concentrations were not always similar, embryo and FG tissues generally showed similar patterns of change in starch and sugar concentrations over time. Total starch concentration was highest during early seed development and decreased as development progressed. The major sugars contributing to osmotic potential during early seed development were D-pinitol, sucrose, fructose and glucose. During mid-seed development, D-pinitol, sucrose, fructose, glucose, melibiose and raffinose provided major contributions to the osmotic environment. During late seed development, sucrose, raffinose, melibiose, stachyose and fructose were the major contributors to osmotic potential. These data suggest stage-specific media composition for each step in the somatic embryogenesis protocol.

  15. Investigating carbohydrate isomers by IMS-CID-IMS-MS: precursor and fragment ion cross-sections. (United States)

    Gaye, M M; Kurulugama, R; Clemmer, D E


    Ion mobility spectrometry techniques (IMS and IMS-IMS) combined with collision-induced dissociation (CID) and mass spectrometry (MS) are used to investigate the structures of singly-lithiated carbohydrate isomers. With the exception of some favorable cases, IMS-MS analyses of underivatized carbohydrates reveal that most isobaric precursor ions have similar collision cross sections (ccs). In contrast, ccs values for isomeric fragment ions obtained by IMS-CID-IMS-MS analysis are often different, and thus appear to be useful as a means of distinguishing the isomeric precursors. We report values of ccs (in He) for precursor- and associated-fragment ions for three monosaccharide isomers (glucose, galactose and fructose), ten disaccharide isomers (sucrose, leucrose, palatinose, trehalose, cellobiose, β-gentiobiose, isomaltose, maltose, lactose and melibiose), and three trisaccharide isomers (raffinose, melezitose and maltotriose). These values are discussed as a means of differentiating precursor carbohydrates.

  16. Physiological studies in aerobic batch cultivations of Saccharomyces cerevisiae strains harboring the MEL1 gene

    DEFF Research Database (Denmark)

    Østergaard, Simon; Roca, Christophe Francois Aime; Ronnow, B.


    Physiological studies of Saccharomyces cerevisiae strains harboring the MEL1 gene were carried out in aerobic batch cultivations on glucose-galactose mixtures and on the disaccharide melibiose, which is hydrolyzed by the enzyme melibiase (Mel1, EC into a glucose and a galactose moiety....... The strains examined (T200, T256, M24, and TH1) were all derived from the bakers' and distillers' strain of S. cerevisiae, DGI 342. All the strains showed a significant higher ethanol yield when growing on glucose, and half the biomass yield, compared with growth on galactose. The maximum specific uptake...... rates were 2.5-3.3-fold higher on glucose than on galactose for all the strains examined, and hence, ethanol production was pronounced on glucose due to respiro-fermentative metabolism. The T256 strain and the T200 strain having the MEL1 gene inserted in the HXK2 locus and the LEU2 locus, respectively...

  17. Transcriptional analysis of oligosaccharide utilization by Bifidobacterium lactis Bl-04

    DEFF Research Database (Denmark)

    Andersen, Joakim Mark; Barrangou, Rodolphe; Abou Hachem, Maher


    BACKGROUND: Probiotic bifidobacteria in combination with prebiotic carbohydrates have documented positive effects on human health regarding gastrointestinal disorders and improved immunity, however the selective routes of uptake remain unknown for most candidate prebiotics. The differential...... of glycoside (galactosides, glucosides or xylosides) utilized. Carbohydrate transporters of the major facilitator superfamily (induced by gentiobiose and β-galacto-oligosaccharides (GOS)) and ATP-binding cassette (ABC) transporters (upregulated by cellobiose, GOS, isomaltose, maltotriose, melibiose, panose......, raffinose, stachyose, xylobiose and β-xylo-oligosaccharides) were differentially upregulated, together with glycoside hydrolases from families 1, 2, 13, 36, 42, 43 and 77. Sequence analysis of the identified solute-binding proteins that determine the specificity of ABC transporters revealed similarities...

  18. MsmK, an ATPase, Contributes to Utilization of Multiple Carbohydrates and Host Colonization of Streptococcus suis. (United States)

    Tan, Mei-Fang; Gao, Ting; Liu, Wan-Quan; Zhang, Chun-Yan; Yang, Xi; Zhu, Jia-Wen; Teng, Mu-Ye; Li, Lu; Zhou, Rui


    Acquisition and metabolism of carbohydrates are essential for host colonization and pathogenesis of bacterial pathogens. Different bacteria can uptake different lines of carbohydrates via ABC transporters, in which ATPase subunits energize the transport though ATP hydrolysis. Some ABC transporters possess their own ATPases, while some share a common ATPase. Here we identified MsmK, an ATPase from Streptococcus suis, an emerging zoonotic bacterium causing dead infections in pigs and humans. Genetic and biochemistry studies revealed that the MsmK was responsible for the utilization of raffinose, melibiose, maltotetraose, glycogen and maltotriose. In infected mice, the msmK-deletion mutant showed significant defects of survival and colonization when compared with its parental and complementary strains. Taken together, MsmK is an ATPase that contributes to multiple carbohydrates utilization and host colonization of S. suis. This study gives new insight into our understanding of the carbohydrates utilization and its relationship to the pathogenesis of this zoonotic pathogen.

  19. Thermoanaerobacter pentosaceus sp. nov., an anaerobic, extreme thermophilic, high ethanol-yielding bacterium isolated from household waste

    DEFF Research Database (Denmark)

    Tomás, Ana Faria; Karakashev, Dimitar Borisov; Angelidaki, Irini


    of approximately 0.5 µm. Optimal growth occurred at 70 °C and pH(25°C) 7, with a maximum growth rate of 0.1 h-1. DNA G+C content was 34.2 mol %. Strain DTU01(T) could ferment arabinose, cellobiose, fructose, galactose, glucose, inulin, lactose, mannose, melibiose, pectin, starch, sucrose, xylan, yeast extract...... and xylose, but not cellulose, Avicel®, mannitol, inositol, glycerol, acetate, lactate, ethanol, butanol or peptone. Ethanol was the major fermentation product and a maximum yield of 1.39 mol of ethanol per mol xylose was achieved when sulphite was added to the cultivation medium. Thiosulphite......, the physiological and phylogenetic differences (DNA G+C content, substrate utilization, electron acceptors, phylogenetic distance, isolation site) allow for the proposal of strain DTU01(T) as a new species within the genus Thermoanaerobacter, for which the name Thermoanaerobacter pentosaceus sp. nov. is proposed...

  20. Thermomyces lanuginosus CBS 395.62/b Strain as Rich Source of α-Galactosidase Enzyme

    Directory of Open Access Journals (Sweden)

    Quang D. Nguyen


    Full Text Available Seventeen Thermomyces lanuginosus strains, cultivated on raffinose and sucrose, were ranked on the basis of α-galactosidase activities. T. lanuginosus CBS 395.62/b strain showed the highest α-galactosidase activity on both investigated carbohydrates. Several carbon sources were tested as potential inducers for the α-galactosidase synthesis. On melibiose substrate α-galactosidase activity was higher in the intracellular fraction than in the filtrate of the fermentation broth, although both values were very low and did not reach the value of 1 U/mL. Raffinose, sucrose and Lactosucrose® proved to be inducers for α-galactosidase production. The highest titer (about 30 U/mL was achieved on 1 % sucrose and 0.45 % ammonium acetate. The optimum sucrose and ammonium acetate concentrations, at which about 90 U/mL α-galactosidase activity was reached during an 8-day fermentation, were 3 and 0.9 %, respectively.

  1. Simultaneous Analysis of Saccharides between Fresh and Processed Radix Rehmanniae by HPLC and UHPLC-LTQ-Orbitrap-MS with Multivariate Statistical Analysis

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    Shujuan Xue


    Full Text Available Radix Rehmanniae (RR is a kind of herb which is widely used in the clinical and food processing industry. There are four forms of RR used in traditional Chinese medicine practice, which include fresh RR (FRR, raw RR (RRR, processed RR (PRR, and another processed RR (APRR, in which the APRR was processed by nine cycles of repeated steaming and drying. There are a large number of saccharides in RR. However, the differences in content were shown by different processing methods. In this study, an effective method using high-performance liquid chromatography (HPLC and high-performance liquid chromatography-mass spectrometry (LC-MS coupled with multivariate statistical analysis to rapidly distinguish different RR samples and validate the proposed chemical conversion mechanism. The datasets of the content of saccharides were subjected to principal component analysis (PCA and one-way analysis of variance. The results showed that there different changes occurred in the contents of saccharides corresponding to the different processing methods, in which the contents of monosaccharides—namely arabinose, glucose, mannose, and galactose—had an increasing trend or remained relatively stable. However, the contents of fructose and oligosaccharides, including manninotriose, melibiose, sucrose, and raffinose, first increased and then reduced, or gradually decreased, yet the content of stachyose gradually decreased. The MSn data indicated that manninotriose, melibiose, and some monosaccharides were produced by the hydrolysis of oligosaccharides. In addition, the fragmentation pathways of 1-phenyl-3-methyl-5-pyrazolone (PMP derivatization of monosaccharides were also found that its glycosidic bond was first broken and subsequently its inside ring broke, and the characteristic fragment ions were produced at m/z 511.22, 493.20, 373.16, and 175.08 in the PMP derivatization of monosaccharides. In conclusion, this study illustrates the change and chemical conversion

  2. A Fungal α-Galactosidase from Tricholoma matsutake with Broad Substrate Specificity and Good Hydrolytic Activity on Raffinose Family Oligosaccharides

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    Xueran Geng


    Full Text Available An acidic α-galactosidase designated as TMG was purified from the fruiting bodies The purification protocol entailed ion exchange chromatography on Q-Sepharose and of Tricholoma matsutake with 136-fold purification and a specific activity of 909 units/mg. Mono-Q and fast protein liquid chromatography on Superdex 75. TMG is a monomeric protein exhibiting a molecular mass of 47 kDa in SDS-PAGE and gel filtration. The purified enzyme was identified by LC-MS/MS and three inner amino acid sequences were obtained. The optimum pH and temperature for TMG with pNPGal as substrate were pH 4.5 and 55 °C, respectively. The α-galactosidase activity was strongly inhibited by K+, Ca2+, Cd2+, Hg2+, Ag+ and Zn2+ ions. The enzyme activity was inhibited by the chemical modification agent N-bromosuccinimide (NBS, indicating the importance of tryptophan residue(s at or near the active site. Besides hydrolyzing pNPGal, TMG also efficaciously catalyzed the degradation of natural substrates such as stachyose, raffinose, and melibiose. Thus TMG can be exploited commercially for improving the nutritional value of soy milk by degradation of indigestible oligosaccharides.

  3. Biochemical and functional properties of a lectin purified from korean large black soybeans--a cultivar of glycine max. (United States)

    Fang, E F; Wong, J H; Lin, P; Ng, T B


    Lectins, a class of proteins that reversibly and non-enzymatically bind specific sugars, have been purified from different kinds of legumes. In this study, a 48-kDa lectin (KBL) was purified from Korean large black soybeans using liquid chromatography. The specific hemagglutinating activity of the KBL was 4096 titer/mg. EDTA-induced loss of hemagglutinating activity of KBL could be recovered by addition of Fe(3+) ions and some divalent cations as Ca(2+), Mn(2+), Fe(2+), Cu(2+), Zn(2+), and Pb(2+). Sugars such as D-(+)-galactose, D-(+)-raffinose, L-(+)-arabinose, alpha-D-(+)-melibiose, and alpha-lactose could inhibit the hemagglutinating activity of the lectin. Furthermore, the protein showed high thermal stability as well as stability over a wide range of pH values. KBL inhibited HIV-1 reverse transcriptase activity with an IC(50) of 1.38 microM. However, it was destitute of cytokine releasing, mitogenic, ribonuclease and antifungal activities. In addition, inhibitory activity toward nasopharyngeal cell lines was undetectable in KBL at concentrations up to 20 microM.

  4. Major facilitator superfamily domain-containing protein 2a (MFSD2A has roles in body growth, motor function, and lipid metabolism.

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    Justin H Berger

    Full Text Available The metabolic adaptations to fasting in the liver are largely controlled by the nuclear hormone receptor peroxisome proliferator-activated receptor alpha (PPARα, where PPARα upregulates genes encoding the biochemical pathway for β-oxidation of fatty acids and ketogenesis. As part of an effort to identify and characterize nutritionally regulated genes that play physiological roles in the adaptation to fasting, we identified Major facilitator superfamily domain-containing protein 2a (Mfsd2a as a fasting-induced gene regulated by both PPARα and glucagon signaling in the liver. MFSD2A is a cell-surface protein homologous to bacterial sodium-melibiose transporters. Hepatic expression and turnover of MFSD2A is acutely regulated by fasting/refeeding, but expression in the brain is constitutive. Relative to wildtype mice, gene-targeted Mfsd2a knockout mice are smaller, leaner, and have decreased serum, liver and brown adipose triglycerides. Mfsd2a knockout mice have normal liver lipid metabolism but increased whole body energy expenditure, likely due to increased β-oxidation in brown adipose tissue and significantly increased voluntary movement, but surprisingly exhibited a form of ataxia. Together, these results indicate that MFSD2A is a nutritionally regulated gene that plays myriad roles in body growth and development, motor function, and lipid metabolism. Moreover, these data suggest that the ligand(s that are transported by MFSD2A play important roles in these physiological processes and await future identification.

  5. Isolation and characterization of novel bacteria containing acc deaminase from the rhizosphere resource on dry-farming lands

    International Nuclear Information System (INIS)

    Zheng, P.; Tian, L.; Chen, F.; Cui, Z.


    Soil-microbe-plant interactions are known to be intricate and they can greatly influence the crop vigor and yield. Plant growth promoting rhizobacteria (PGPR) containing ACC deaminase can markedly affect plant metabolic processes under stress conditions. In the present study, we isolated 300 bacterial strains from the rhizosphere of maize or apple grown in drought-hit soil including four different locations of the Loess Plateau, China. Of all isolated strains, four with ACC deaminase activity (ranging from 28.88 to 155.12 nmol alpha-ketobutyrate mg-1 h-1) were further studied by determining their biological characters and sequencing the 16S rRNA gene. All four strains showed positive performance in terms of arabinose, citrate utilization, urease, indol, glucose and melibiose. In connection with the results of biochemical characters and phylogenetic analysis, these strains commonly belong to three different genera: Klebsiella, Pseudomonas and Raoultella and four different species: Klebsiella oxytoca, Klebsiella variicola, Pseudomonas fluorescens and Raoultella planticola. Although some researchers have reported their performance under stress conditions, we are the first to report Klebsiella oxytoca, Klebsiella variicola and Raoultella planticola containing ACC deaminase under drought stress. These findings are a reasonable explanation to their superb ability of causing stress-resistance in maize (Zea mays) or apple (Malus domestica) plants. The presence of diverse PGPR possessing potential ACC deaminase activity may be beneficial for enhancing crop production under different stress conditions. (author)

  6. Reclassification of Lactobacillus kimchii and Lactobacillus bobalius as later subjective synonyms of Lactobacillus paralimentarius. (United States)

    Pang, Huili; Kitahara, Maki; Tan, Zhongfang; Wang, Yanping; Qin, Guangyong; Ohkuma, Moriya; Cai, Yimin


    Characterization and identification of strain CW 1 ( = JCM 17161) isolated from corn silage were performed. Strain CW 1 was a Gram-positive, catalase-negative and homofermentative rod that produced the DL-form of lactic acid. This strain exhibited more than 99.6% 16S rRNA gene sequence similarity and greater than 82% DNA-DNA reassociation with type strains of Lactobacillus kimchii, L. bobalius and L. paralimentarius. To clarify the taxonomic positions of these type strains, phenotypic characterization, 16S rRNA gene sequencing, ribotyping and DNA-DNA relatedness were examined. The three type strains displayed different L-arabinose, lactose, melibiose, melezitose, raffinose and N-acetyl-β-glucosaminidase fermentation patterns. Phylogenetic analysis showed that L. paralimentarius is a closer neighbour of L. kimchii and L. bobalius, sharing 99.5-99.9% 16S rRNA gene sequence similarity, which was confirmed by the high DNA-DNA relatedness (≥82%) between L. paralimentarius JCM 10415(T), L. bobalius JCM 16180(T) and L. kimchii JCM 10707(T). Therefore, it is proposed that L. kimchii and L. bobalius should be reclassified as later synonyms of L. paralimentarius.

  7. [Comparison of efficacy of tests for differentiation of typical and atypical strains of Yersinia pestis and Yersinia pseudotuberculosis]. (United States)

    Arsen'eva, T E; Lebedeva, S A; Trukhachev, A L; Vasil'eva, E A; Ivanova, V S; Bozhko, N V


    To characterize species specificity of officially recommended tests for differentiation of Yersiniapestis and Yersinia pseudotuberculosis and propose additional tests allowing for more accurate identification. Natural, laboratory and typical strains oftwo Yersinia species were studied using microbiological, molecular and biochemical methods. For PCR species-specific primers complementary to certain fragments of chromosomal DNA of each species as well as to several plasmid genes of Y. pestis were used. It was shown that such attributes of Y. pestis as form of colonies, fermentation ofrhamnose, melibiose and urea, susceptibility to diagnostic phages, nutritional requirements could be lost in pestis bacterial species or detected in pseudotuberculosis species. Such attribute as mobility as well as positive result of CoA-reaction on fraction V antigen are more reliable. Guaranteed differentiation of typical and changed according to differential tests strains is provided only by PCR-analysis with primers vlml2for/ISrev216 and JS respectively, which are homologous to certain chromosome fragments of one of two Yersinia species.

  8. Involvement of L(-)-rhamnose in sea urchin gastrulation: a live embryo assay. (United States)

    Smith, Tiffany N; Oppenheimer, Steven B


    The sea urchin embryo is a National Institutes of Health model system that has provided major developments, and is important in human health and disease. To obtain initial insights to identify glycans that mediate cellular interactions, Lytechinus pictus sea urchin embryos were incubated at 24 or 30 h post-fertilization with 0.0009-0.03 M alpha-cyclodextrin, melibiose, L(-)-rhamnose, trehalose, D(+)-xylose or L(-)-xylose in lower-calcium artificial sea water (pH 8.0, 15°C), which speeds the entry of molecules into the interior of the embryos. While α-cyclodextrin killed the embryos, and L(-)-xylose had small effects at one concentration tested, L(-)-rhamnose caused substantially increased numbers of unattached archenterons and exogastrulated embryos at low glycan concentrations after 18-24 h incubation with the sugar. The results were statistically significant compared with the control embryos in the absence of sugar (P embryos were examined. This study is the first demonstration that uses a live embryo assay for a likely role for L(-)-rhamnose in sea urchin gastrula cellular interactions, which have interested investigators for over a century.

  9. Antiproliferative effects of galectin-1 from Rana catesbeiana eggs on human leukemia cells and its binding proteins in human cells. (United States)

    Yasumitsu, Hidetaro; Mochida, Keiichi; Yasuda, Chie; Isobe, Masaharu; Kawsar, Sarkar M A; Fujii, Yuki; Matsumoto, Ryo; Kanaly, Robert A; Ozeki, Yasuhiro


    Galectin-1 from American bullfrog, RCG1, was isolated to high purity, and its growth inhibitory properties against human cells were examined. The results demonstrated that highly purified RCG1 induced large cell aggregates and revealed cell-type-specific growth inhibition. It significantly inhibited all human leukemia cell lines tested such as HL-60, U937, and K562 cells but did not inhibit human colon cancer cell line, Colo 201, or mouse mammary tumor cell line FM3A cells. Although most of the galectin-induced growth inhibitions are known to be apoptic, RCG1 induced growth arrest and neither apoptosis nor necrosis. RCG1-mediated growth inhibition was specifically suppressed by the corresponding sugar, lactose, but not by sucrose or even the structurally similar sugar, melibiose. Several studies have reported that galectin-mediated biological functions were modulated by charge modification. Since the high purity of RCG1 was demonstrated but a moderate degree of growth inhibition occurred, it is possible protein charge modification was examined by isoelectric focusing, and it was found to be highly heterogeneous in charge. RCG1 binding proteins in human cells were analyzed by lectin blotting using biotinylated RCG1, and lectin blotting revealed that in human cell extracts the specific proteins at molecular weight 37 and 50 kDa possessed the responsive features of RCG1 binding and lactose competition.

  10. Atypical Yersinia pseudotuberculosis serotype O:3 isolated from hunted wild boars in Italy. (United States)

    Magistrali, C F; Cucco, L; Manuali, E; Sebastiani, C; Farneti, S; Ercoli, L; Pezzotti, G


    Atypical Yersinia pseudotuberculosis serotype O:3 was isolated from rectal contents of two wild boars hunted in Italy within a regional wildlife management program. No outbreak of yersiniosis was reported in this area in the same period and no lesions were found by the veterinarian at post-mortem inspection. Nevertheless, after histological examination, granulomatous lesions were detected in submandibular lymph nodes of one of the two wild boars. Microbiological and bio molecular characterization of the isolates revealed a melibiose-negative, biotype 2, wbyK+O:3 genotype, carrying inv, yop (yopH and yopB), virF, and R-HPI. Strains showing the same profile, matching to the criteria of genetic group 5, have been recently reported in fatal cases of yersiniosis in cynomolgus macaques and in farmed deer and atypical O:3 serotype has been suggested as a pathogenic subtype of O:3. This is the third report of an atypical O:3 Y. pseudotuberculosis strain, the first outside the American continent and the first one not associated to fatal yersiniosis. Wild boars could be a possible reservoir of this emerging pathogen. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Isolation from a shea cake digester of a tannin-tolerant Escherichia coli strain decarboxylating p-hydroxybenzoic and vanillic acids. (United States)

    Chamkha, Mohamed; Record, Eric; Garcia, Jean-Louis; Asther, Marcel; Labat, Marc


    A facultatively anaerobic, mesophilic, Gram-negative, non-motile, non-sporulated bacterium, designated strain C2, was isolated from an anaerobic digester fed with shea cake rich in tannins and aromatic compounds and previously inoculated with anaerobic sludge from the pit of a slaughterhouse, after enrichment on tannic acid. The straight rods occurred singly or in pairs. Strain C2 fermented numerous carbohydrates (fructose, galactose, glucose, lactose, mannose, maltose, melibiose, raffinose, rhamnose, ribose, saccharose, sorbitol, trehalose, and xylose) and peptides (Biotrypcase, Casamino acids, and yeast extract), producing acid and gas, and had a G + C content of 51.6 +/- 0.1 mol %. Strain C2 was very closely related to Escherichia coli (= DSM 30083(T)) phylogenetically (similarity of 99%), genotypically (DNA homology of 79%), and phenotypically. The isolate tolerated tannic acid (hydrolyzable tannin) and decarboxylated by non-oxidative decarboxylation only p-hydroxybenzoic and vanillic acids to their corresponding phenol and guaicol, under anaerobic and aerobic conditions without further degradation. Adding glucose increased growth and the rate of conversion. High concentrations of p-hydroxybenzoic acid or vanillic acid inhibited growth, and decarboxylation could not occur completely, suggesting phenol toxicity. In contrast, the type strain of E. coli cannot metabolize p-hydroxybenzoic and vanillic acids, anaerobically or aerobically, with or without glucose added.

  12. Partial Characterization of α-Galactosidic Activity from the Antarctic Bacterial Isolate, . LX-20 as a Potential Feed Enzyme Source

    Directory of Open Access Journals (Sweden)

    Inkyung Park


    Full Text Available An Antarctic bacterial isolate displaying extracellular α-galactosidic activity was named Paenibacillus sp. LX-20 based on 16S rRNA gene sequence analysis. Optimal activity for the LX-20 α-galactosidase occurred at pH 6.0–6.5 and 45°C. The enzyme immobilized on the smart polymer Eudragit L-100 retained 70% of its original activity after incubation for 30 min at 50°C, while the free enzyme retained 58% of activity. The enzyme had relatively high specificity for α-D-galactosides such as p-nitrophenyl-α-galactopyranoside, melibiose, raffinose and stachyose, and was resistant to some proteases such as trypsin, pancreatin and pronase. Enzyme activity was almost completely inhibited by Ag+, Hg2+, Cu2+, and sodium dodecyl sulfate, but activity was not affected by β-mercaptoethanol or EDTA. LX-20 α-galactosidase may be potentially useful as an additive for soybean processing in the feed industry.

  13. Characterization of the plant growth promoting bacterium, Enterobacter cloacae MSR1, isolated from roots of non-nodulating Medicago sativa. (United States)

    Khalifa, Ashraf Y Z; Alsyeeh, Abdel-Moneium; Almalki, Mohammed A; Saleh, Farag A


    The aim of the present study was to characterize the endophytic bacterial strain designated MSR1 that was isolated from inside the non-nodulating roots of Medicago sativa after surface-sterilization. MSR1 was identified as Enterobacter cloacae using both 16S rDNA gene sequence analysis and API20E biochemical identification system (Biomerieux, France). Furthermore, this bacterium was characterized using API50CH kit (Biomerieux, France) and tested for antibacterial activities against some food borne pathogens. The results showed that E. cloacae consumed certain carbohydrates such as glycerol, d-xylose, d-maltose and esculin melibiose as a sole carbon source and certain amino acids such as arginine, tryptophan ornithine as nitrogen source. Furthermore, MSR1 possessed multiple plant-growth promoting characteristics; phosphate solubility, production of phytohormones acetoin and bioactive compounds. Inoculation of Pisum sativum with MSR1 significantly improved the growth parameters (the length and dry weight) of this economically important grain legume compared to the non-treated plants. To our knowledge, this is the first report addressing E. cloacae which exist in roots of alfalfa growing in Al-Ahsaa region. The results confirmed that E. cloacae exhibited traits for plant growth promoting and could be developed as an eco-friendly biofertilizer for P. sativum and probably for other important plant species in future.

  14. Crystal structure of the enzyme-product complex reveals sugar ring distortion during catalysis by family 63 inverting α-glycosidase. (United States)

    Miyazaki, Takatsugu; Nishikawa, Atsushi; Tonozuka, Takashi


    Glycoside hydrolases are divided into two groups, known as inverting and retaining enzymes, based on their hydrolytic mechanisms. Glycoside hydrolase family 63 (GH63) is composed of inverting α-glycosidases, which act mainly on α-glucosides. We previously found that Escherichia coli GH63 enzyme, YgjK, can hydrolyze 2-O-α-d-glucosyl-d-galactose. Two constructed glycosynthase mutants, D324N and E727A, which catalyze the transfer of a β-glucosyl fluoride donor to galactose, lactose, and melibiose. Here, we determined the crystal structures of D324N and E727A soaked with a mixture of glucose and lactose at 1.8- and 2.1-Å resolutions, respectively. Because glucose and lactose molecules are found at the active sites in both structures, it is possible that these structures mimic the enzyme-product complex of YgjK. A glucose molecule found at subsite -1 in both structures adopts an unusual 1 S 3 skew-boat conformation. Comparison between these structures and the previously determined enzyme-substrate complex structure reveals that the glucose pyranose ring might be distorted immediately after nucleophilic attack by a water molecule. These structures represent the first enzyme-product complex for the GH63 family, as well as the structurally-related glycosidases, and it may provide insight into the catalytic mechanism of these enzymes. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Structure of a bacterial glycoside hydrolase family 63 enzyme in complex with its glycosynthase product, and insights into the substrate specificity. (United States)

    Miyazaki, Takatsugu; Ichikawa, Megumi; Yokoi, Gaku; Kitaoka, Motomitsu; Mori, Haruhide; Kitano, Yoshikazu; Nishikawa, Atsushi; Tonozuka, Takashi


    Proteins belonging to glycoside hydrolase family 63 (GH63) are found in bacteria, archaea and eukaryotes. Although the eukaryotic GH63 proteins have been identified as processing α-glucosidase I, the substrate specificities of the bacterial and archaeal GH63 proteins are not clear. Here, we converted a bacterial GH63 enzyme, Escherichia coli YgjK, to a glycosynthase to probe its substrate specificity. Two mutants of YgjK (E727A and D324N) were constructed, and both mutants showed glycosynthase activity. The reactions of E727A with β-D-glucosyl fluoride and monosaccharides showed that the largest amount of glycosynthase product accumulated when galactose was employed as an acceptor molecule. The crystal structure of E727A complexed with the reaction product indicated that the disaccharide bound at the active site was 2-O-α-D-glucopyranosyl-α-D-galactopyranose (Glc12Gal). A comparison of the structures of E727A-Glc12Gal and D324N-melibiose showed that there were two main types of conformation: the open and closed forms. The structure of YgjK adopted the closed form when subsite -1 was occupied by glucose. These results suggest that sugars containing the Glc12Gal structure are the most likely candidates for natural substrates of YgjK. © 2013 FEBS.

  16. Aspergillus nidulans α-galactosidase of glycoside hydrolase family 36 catalyses the formation of α-galacto-oligosaccharides by transglycosylation

    DEFF Research Database (Denmark)

    Nakai, Hiroyuki; Baumann, Martin; Petersen, B. O.


    The α-galactosidase from Aspergillus nidulans (AglC) belongs to a phylogenetic cluster containing eukaryotic α-galactosidases and α-galacto-oligosaccharide synthases of glycoside hydrolase family 36 (GH36). The recombinant AglC, produced in high yield (0.65 g·L−1 culture) as His-tag fusion...... in Escherichia coli, catalysed efficient transglycosylation with α-(1→6) regioselectivity from 40 mm 4-nitrophenol α-d-galactopyranoside, melibiose or raffinose, resulting in a 37–74% yield of 4-nitrophenol α-d-Galp-(1→6)-d-Galp, α-d-Galp-(1→6)-α-d-Galp-(1→6)-d-Glcp and α-d-Galp-(1→6)-α-d-Galp-(1→6)-d-Glcp-(α1→β......2)-d-Fruf (stachyose), respectively. Furthermore, among 10 monosaccharide acceptor candidates (400 mm) and the donor 4-nitrophenol α-d-galactopyranoside (40 mm), α-(1→6) linked galactodisaccharides were also obtained with galactose, glucose and mannose in high yields of 39–58%. AglC did...

  17. The Carbohydrate assimilation pattern in Iranian typical and atypical strains of Microsporum Canis

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    Zaini F


    Full Text Available The values of fourteen carbohydrates assimilation patterns were investigated for typical and atypical strains of Microsporum canis. Thiry eight strains of typical and twenty two strains of atypical Microsporum canis, Microsporum canis NCPF 352 and one Microsporum distortum were included in this study. Statistical analysis of the results indicated that despite limited variations within the pattern of carbohydrate utilization, no correlation. The results also revealed that erythritol and trehalose were best utilized for sporulation by the typical and atypical strains of Microsporum canis. Production of obundant macroconidia, microconidia and chlamydoconidia by use of erythritol and trehalose suggested that these two carbohydrates were effective in production of fluffly appearance in colonies examined. The Microsporum canis NCPF 352 strongly utilized glucose, mannitol and melibiose in addition to the two above-mentioned carbohydrates. Weak erythritol assimilation was observed by Microsporum distortum.Carbohydrate utilization pattern is unable to differentiate typical and atypical strains of Microsporum canis. But it could be regarded as a valuable aid for identification of Microsporum distortum as well as marker in epidemiological investigations.

  18. Identification, technological and safety characterization of Lactobacillus sakei and Lactobacillus curvatus isolated from Argentinean anchovies (Engraulis anchoita). (United States)

    Belfiore, Carolina; Raya, Raúl R; Vignolo, Graciela M


    In this study, the identification and characterization of Lactobacillus previously isolated from fresh anchovies (Engraulis anchoita) are investigated. 16S rDNA partial sequencing assigned all the isolates to belong to the Lactobacillus sakei/curvatus group. Fourteen out of 15 isolates were identified as L. sakei by phenotypic traits: they exhibited catalase activity and fermented melibiose, although only 10 of them hydrolyzed arginine. These results were confirmed by multiplex PCR-based restriction enzyme analysis with HindIII and by restriction fragment length polymorphic (RFLP) analysis of the 16S-23S rDNA intergenic spacer region with TaqI. Among identified isolates, four L. sakei strains and the sole L. curvatus strain showing sensitivity to chloramphenicol, erythromycin and tetracycline and exhibiting high tolerance to NaCl (10-18%) were unable to produce neither dextran nor biogenic amines. Based on technological and safety features, L. sakei SACB704 and L. curvatus SACB03a naturally present in fresh anchovies may be promising strains for the development of a starter culture to accelerate and control the fermentation of salt fermented anchovy-based products.

  19. Production of α-galactosidase from Aspergillus foetidus MTCC 6322 by solid state fermentation and its application in soymilk hydrolysis. (United States)

    Boopathy, Naidu Ramachandra; Gupta, Rishikesh Kumar; Ramudu, Kamini Numbi


    The production of α-galactosidase from the wild fungal strain Aspergillus foetidus MTCC 6322 using solid state fermentation (SSF), its characterization, and its efficacy in the hydrolysis of soymilk using response surface methodology were studied. The optimum conditions for production of α-galactosidase by SSF were: wheat bran (10 g), moisture content (64%), inoculum volume (1.0 mL; 6 x 10(7) spores/mL) with a yield of 4.1 x 10(3) units per gram dry substrate (U/gds) at 96 h. The enzyme showed optimum activity at 6.0, temperature 40 degrees C, pH stability between 5.0-8.0, and temperature stability between 30-40 degrees C. The enzyme was stable in the presence of trypsin, lipase, and collagenase and it showed susceptibility of the substrates such as raffinose, melibiose, guar gum and soymilk to hydrolysis in varying degrees. The optimized conditions for soymilk hydrolysis were: soymilk (10 mL) from defatted soybean meal (1.5%), α-galactosidase (0.15 UmL(-1) at 30 degrees C, pH 6.0 and duration of 1 h.

  20. Caracterização de alfagalactosidase e sua relação com a germinação das sementes de Caesalpinia peltophoroides (Leguminosae Caesalpinioideae Alphagalactosidase characterization and its relationship with seed germination of Caesalpinia peltophoroides (Legumonosae Caesalpinioideae

    Directory of Open Access Journals (Sweden)

    Eduardo Euclydes de Lima e Borges


    Full Text Available Sementes de Caesalpinia peltophoroides (Leguminosae Caesalpinioideae - sibipiruna - foram colocadas para embebição por 144 h, sendo retiradas amostras para análise de proteína, quantificações da atividade de alfagalactosidase e de açúcares presentes na micrópila. A germinação iniciou-se com 96 h de embebição, sem que fossem detectadas modificações na parede celular da micrópila. Nesta, observou-se maior proporção de arabinose, que mostrou tendência de aumento com o decorrer da embebição. A atividade específica da alfagalactosidase foi detectada em sementes secas, tanto no eixo embrionário quanto nos cotilédones, aumentando no primeiro a partir de 24 h de embebição. O aumento da atividade nos cotilédones foi mais lento, sendo mais acentuado a partir de 120 h de embebição. O teor de proteína decresceu continuamente no eixo embrionário a partir de 24 horas de embebição, enquanto se manteve estável nos cotilédones. A atividade da alfagalactosidase foi máxima nas temperaturas de 55 e 50 ºC para o eixo embrionário e para os cotilédones, respectivamente. O pH que mais estimulou a atividade da enzima foi na faixa de 5,5 a 6,0 para o eixo embrionário e na de 4,5 a 5,0 para os cotilédones. As alfagalactosidase do eixo embrionário e dos cotilédones foram inibidas por SDS, CuSO4, galactose e melibiose. Não houve efeito estimulante sobre a atividade da alfagalactosidase do eixo embrionário por nenhum dos efetores, enquanto o mercaptoetanol estimulou a atividade da enzima dos cotilédones. Os K M para o substrato ro-NPGal para a alfagalactosidase do eixo embrionário e dos cotilédones foram de 1,74 e 2,64 mM, respectivamente.Caesalpinia peltophoroides (Leguminosae Caesalpinioideae seeds were soaked in water for 144 hours. Samples were taken for protein analysis, quantification of alphagalactosidase activity and micropyle sugar composition. Germination began after 96 hours of imbibition, with no modifications in

  1. Effector Overlap between the lac and mel Operons of Escherichia coli: Induction of the mel Operon with β-Galactosides (United States)

    Narang, Atul


    ABSTRACT The lac (lactose) operon (which processes β-galactosides) and the mel (melibiose) operon (which processes α-galactosides) of Escherichia coli have a close historical connection. A number of shared substrates and effectors of the permeases and regulatory proteins have been reported over the years. Until now, β-thiogalactosides like TMG (methyl-β-d-thiogalactopyranoside) and IPTG (isopropyl-β-d-thiogalactopyranoside) have not generally been considered to be inducers of the mel operon. The same is true for β-galactosides such as lactose [β-d-galactopyranosyl-(1→4)-d-glucose], which is a substrate but is not itself an inducer of the lac operon. This report shows that all three sugars can induce the mel operon significantly when they are accumulated in the cell by Lac permease. Strong induction by β-thiogalactosides is observed in the presence of Lac permease, and strong induction by lactose (more than 200-fold) is observed in the absence of β-galactosidase. This finding calls for reevaluation of TMG uptake experiments as assays for Lac permease that were performed with mel+ strains. IMPORTANCE The typical textbook picture of bacterial operons is that of stand-alone units of genetic information that perform, in a regulated manner, well-defined cellular functions. Less attention is given to the extensive interactions that can be found between operons. Well-described examples of such interactions are the effector molecules shared by the lac and mel operons. Here, we show that this set has to be extended to include β-galactosides, which have been, until now, considered not to effect the expression of the mel operon. That they can be inducers of the mel operon as well as the lac operon has not been noted in decades of research because of the Escherichia coli genetic background used in previous studies. PMID:28193904

  2. Lactose metabolism in Lactobacillus bulgaricus: analysis of the primary structure and expression of the genes involved. (United States)

    Leong-Morgenthaler, P; Zwahlen, M C; Hottinger, H


    The genes coding for the lactose permease and beta-galactosidase, two proteins involved in the metabolism of lactose by Lactobacillus bulgaricus, have been cloned, expressed, and found functional in Escherichia coli. The nucleotide sequences of these genes and their flanking regions have been determined, showing the presence of two contiguous open reading frames (ORFs). One of these ORFs codes for the lactose permease gene, and the other codes for the beta-galactosidase gene. The lactose permease gene is located in front of the beta-galactosidase gene, with 3 bp in the intergenic region. The two genes are probably transcribed as one operon. Primer extension studies have mapped a promoter upstream from the lactose permease gene but not the beta-galactosidase gene. This promoter is similar to those found in E. coli with general characteristics of GC-rich organisms. In addition, the sequences around the promoter contain a significantly higher number of AT base pairs (80%) than does the overall L. bulgaricus genome, which is rich in GC (GC content of 54%). The amino acid sequences obtained from translation of the ORFs are found to be highly homologous (similarity of 75%) to those from Streptococcus thermophilus. The first 460 amino acids of the lactose permease shows homology to the melibiose transport protein of E. coli. Little homology was found between the lactose permease of L. bulgaricus and E. coli, but the residues which are involved in the binding and the transport of lactose are conserved. The carboxy terminus is similar to that of the enzyme III of several phosphoenolpyruvate-dependent phosphotransferase systems. Images PMID:1705929

  3. Lactobacillus herbarum sp. nov., a species related to Lactobacillus plantarum. (United States)

    Mao, Yuejian; Chen, Meng; Horvath, Philippe


    Strain TCF032-E4 was isolated from a traditional Chinese fermented radish. It shares >99% 16S rRNA sequence identity with L. plantarum, L. pentosus and L. paraplantarum. This strain can ferment ribose, galactose, glucose, fructose, mannose, mannitol, N-acetylglucosamine, amygdalin, arbutin, salicin, cellobiose, maltose, lactose, melibiose, trehalose and gentiobiose. It cannot ferment sucrose, which can be used by L. pentosus, L. paraplantarum, L. fabifermentans, L. xiangfangensis and L. mudanjiangensis, as well as most of the L. plantarum strains (88.7%). TCF032-E4 cannot grow at temperature above 32 °C. This strain shares 78.2-83.6% pheS (phenylalanyl-tRNA synthetase alpha subunit) and 89.5-94.9% rpoA (RNA polymerase alpha subunit) sequence identity with L. plantarum, L. pentosus, L. paraplantarum, L. fabifermentans, L. xiangfangensis and L. mudanjiangensis. These results indicate that TCF032-E4 represents a distinct species. This hypothesis was further confirmed by whole-genome sequencing and comparison with available genomes of related species. The draft genome size of TCF032-E4 is approximately 2.9 Mb, with a DNA G+C content of 43.5 mol%. The average nucleotide identity (ANI) between TCF032-E4 and related species ranges from 79.0 to 81.1%, the highest ANI value being observed with L. plantarum subsp. plantarum ATCC 14917T. A novel species, Lactobacillus herbarum sp. nov., is proposed with TCF032-E4T ( = CCTCC AB2015090T = DSM 100358T) as the type strain.

  4. Characterization of α-galacto-oligosaccharides formed via heterologous expression of α-galactosidases from Lactobacillus reuteri in Lactococcus lactis. (United States)

    Wang, Yvonne; Black, Brenna A; Curtis, Jonathan M; Gänzle, Michael G


    α-Galacto-oligosaccharides (α-GOS) are produced by transgalactosylation reactions of α-galactosidase (α-Gal) or by conversion of raffinose family oligosaccharides by levansucrase. Similarly to β-GOS, α-GOS have the potential to mimic glycan receptors on eukaryotic cells and act as molecular decoys to prevent bacterial infection; however, data on transgalactosylation reactions of α-Gal remain scarce. The α-Gal gene sequence from Lactobacillus reuteri was cloned into an α-Gal negative strain of Lactococcus lactis. Transgalactosylation reactions were achieved using crude cell extracts with melibiose or raffinose as galactosyl donor and fucose, N-acetylglucosamine or lactose as galactosyl acceptor. The composition, sequence and most linkage types of α-GOS formed with acceptors saccharides were determined by liquid chromatography-tandem mass spectrometry. α-Gal of Lactobacillus reuteri formed (1 → 3)-, (1 → 4)- or (1 → 6)-linked α-GOS but exhibited a preference for formation of (1 → 6)-linkages. Fucose, N-acetylglucosamine and lactose were suitable galactosyl acceptors for α-Gal of L. reuteri, resulting in formation of (1 → 3)-, (1 → 4)- or (1 → 6)-linked hetero-oligosaccharides. By determining the structural specificity of α-Gal and increasing the variation of oligosaccharides produced by introducing alternative acceptor sugars, this work supports further studies to assess α-GOS pathogen adhesion prevention in mammalian hosts.

  5. The new species Enterobacter oryziphilus sp. nov. and Enterobacter oryzendophyticus sp. nov. are key inhabitants of the endosphere of rice (United States)


    Background Six independent Gram-negative, facultatively anaerobic, non-spore-forming, nitrogen-fixing rod-shaped isolates were obtained from the root endosphere of rice grown at the International Rice Research Institute (IRRI) and investigated in a polyphasic taxonomic study. Results The strains produced fatty acid patterns typical for members of the family Enterobacteriaceae. Comparative sequence analyses of the 16S rRNA as well as rpoB genes allocated the strains to two well-defined groups within the genus Enterobacter, family Enterobacteriaceae. The analyses indicated Enterobacter radicincitans, Enterobacter arachidis and Enterobacter oryzae to be the closest related species. An RpoB (translated) protein comparison supported the placement in the genus Enterobacter and the relatedness of our isolates to the aforementioned species. Genomic DNA:DNA hybridization analyses and biochemical analyses provided further evidence that the novel strains belong to two new species within the genus Enterobacter. The two species can be differentiated from each other and from existing enteric species by acid production from L-rhamnose and D-melibiose, decarboxylation of ornithine and utilization of D-alanine, D-raffinose L-proline and L-aspartic acid, among other characteristics. Members of both species revealed capacities to colonise rice roots, including plant-growth-promoting capabilities such as an active supply of fixed nitrogen to the plant and solubilisation of inorganic phosphorus, next to traits allowing adaptation to the plant. Conclusions Two novel proposed enterobacterial species, denominated Enterobacter oryziphilus sp. nov. (type strain REICA_142T=LMG 26429T=NCCB 100393T) and Enterobacter oryzendophyticus sp. nov. (type strain REICA_082T=LMG 26432T =NCCB 100390T) were isolated from rice roots. Both species are capable of promoting rice growth by supplying nitrogen and phosphorus. PMID:23865888

  6. Photobacterium jeanii sp. nov., isolated from corals and zoanthids. (United States)

    Chimetto, Luciane A; Cleenwerck, Ilse; Thompson, Cristiane C; Brocchi, Marcelo; Willems, Anne; De Vos, Paul; Thompson, Fabiano L


    Four novel isolates (R-40508(T), R-40507, R-40903 and R-21419) were obtained from different cnidarian species (Phyllogorgia dilatata, Merulina ampliata and Palythoa caribaeorum) from different places in Brazil and Australia. The novel isolates formed a tight phylogenetic group based on 16S rRNA, recA, topA, ftsZ, mreB and rpoA gene sequences. Their closest phylogenetic neighbours were the type strains of Photobacterium leiognathi, P. rosenbergii and P. halotolerans, sharing 97.1-97.5 % 16S rRNA gene sequence similarity. DNA-DNA hybridization between a representative strain (R-40508(T)) and the type strains of these Photobacterium species revealed less than 20 % relatedness, showing that the new isolates belong to a novel species. Several phenotypic features allow the differentiation of the novel species from its closest phylogenetic neighbours. It has gelatinase and lipase activity and can utilize melibiose, but it cannot grow on 6 % NaCl. In addition, the novel species has the fatty acid iso-C(16 : 0), but lacks the fatty acids C(17 : 0), C(17 : 0) cyclo, iso-C(17 : 0), C(17 : 1)ω8c and iso-C(17 : 1)ω9c. The name Photobacterium jeanii sp. nov. is proposed for this species, with the type strain R-40508(T) (=LMG 25436(T) =CAIM 1817(T)). The G+C content of the type strain is 45.5mol%.

  7. Effector Overlap between the lac and mel Operons of Escherichia coli: Induction of the mel Operon with β-Galactosides. (United States)

    Narang, Atul; Oehler, Stefan


    The lac (lactose) operon (which processes β-galactosides) and the mel (melibiose) operon (which processes α-galactosides) of Escherichia coli have a close historical connection. A number of shared substrates and effectors of the permeases and regulatory proteins have been reported over the years. Until now, β-thiogalactosides like TMG (methyl-β-d-thiogalactopyranoside) and IPTG (isopropyl-β-d-thiogalactopyranoside) have not generally been considered to be inducers of the mel operon. The same is true for β-galactosides such as lactose [β-d-galactopyranosyl-(1→4)-d-glucose], which is a substrate but is not itself an inducer of the lac operon. This report shows that all three sugars can induce the mel operon significantly when they are accumulated in the cell by Lac permease. Strong induction by β-thiogalactosides is observed in the presence of Lac permease, and strong induction by lactose (more than 200-fold) is observed in the absence of β-galactosidase. This finding calls for reevaluation of TMG uptake experiments as assays for Lac permease that were performed with mel + strains. IMPORTANCE The typical textbook picture of bacterial operons is that of stand-alone units of genetic information that perform, in a regulated manner, well-defined cellular functions. Less attention is given to the extensive interactions that can be found between operons. Well-described examples of such interactions are the effector molecules shared by the lac and mel operons. Here, we show that this set has to be extended to include β-galactosides, which have been, until now, considered not to effect the expression of the mel operon. That they can be inducers of the mel operon as well as the lac operon has not been noted in decades of research because of the Escherichia coli genetic background used in previous studies. Copyright © 2017 American Society for Microbiology.

  8. Water deficit affects primary metabolism differently in two Lolium multiflorum/Festuca arundinacea introgression forms with a distinct capacity for photosynthesis and membrane regeneration.

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    Dawid Perlikowski


    Full Text Available Understanding how plants respond to drought at different levels of cell metabolism is an important aspect of research on the mechanisms involved in stress tolerance. Furthermore, a dissection of drought tolerance into its crucial components by the use of plant introgression forms facilitates to analyze this trait more deeply. The important components of plant drought tolerance are the capacity for photosynthesis under drought conditions, and the ability of cellular membrane regeneration after stress cessation. Two closely related introgression forms of Lolium multiflorum/Festuca arundinacea, differing in the level of photosynthetic capacity during stress, and in the ability to regenerate their cellular membranes after stress cessation, were used as forage grass models in a primary metabolome profiling and in an evaluation of chloroplast 1,6-bisphosphate aldolase accumulation level and activity, during 11 days of water deficit, followed by 10 days of rehydration. It was revealed here that the introgression form, characterized by the ability to regenerate membranes after rehydration, contained higher amounts of proline, melibiose, galactaric acid, myo-inositol and myo-inositol-1-phosphate involved in osmoprotection and stress signaling under drought. Moreover, during the rehydration period, this form also maintained elevated accumulation levels of most the primary metabolites, analyzed here. The other introgression form, characterized by the higher capacity for photosynthesis, revealed a higher accumulation level and activity of chloroplast aldolase under drought conditions, and higher accumulation levels of most photosynthetic products during control and drought periods. The potential impact of the observed metabolic alterations on cellular membrane recovery after stress cessation, and on a photosynthetic capacity under drought conditions in grasses, are discussed.

  9. Caldicoprobacter algeriensis sp. nov. a new thermophilic anaerobic, xylanolytic bacterium isolated from an Algerian hot spring. (United States)

    Bouanane-Darenfed, Amel; Fardeau, Marie-Laure; Grégoire, Patrick; Joseph, Manon; Kebbouche-Gana, Salima; Benayad, Tahar; Hacene, Hocine; Cayol, Jean-Luc; Ollivier, Bernard


    A thermophilic anaerobic bacterium (strain TH7C1(T)) was isolated from the hydrothermal hot spring of Guelma in the northeast of Algeria. Strain TH7C1(T) stained Gram-positive, was a non-motile rod appearing singly, in pairs, or as long chains (0.7-1 × 2-6 μm(2)). Spores were never observed. It grew at temperatures between 55 and 75°C (optimum 65°C) and at pH between 6.2 and 8.3 (optimum 6.9). It did not require NaCl for growth, but tolerated it up to 5 g l(-1). Strain TH7C1(T) is an obligatory heterotroph fermenting sugars including glucose, galactose, lactose, raffinose, fructose, ribose, xylose, arabinose, maltose, mannitol, cellobiose, mannose, melibiose, saccharose, but also xylan, and pyruvate. Fermentation of sugars only occurred in the presence of yeast extract (0.1%). The end-products from glucose fermentation were acetate, lactate, ethanol, CO(2), and H(2). Nitrate, nitrite, thiosulfate, elemental sulfur, sulfate, and sulfite were not used as electron acceptors. The G+C content of the genomic DNA was 44.7 mol% (HPLC techniques). Phylogenetic analysis of the small-subunit ribosomal RNA (rRNA) gene sequence indicated that strain TH7C1(T) was affiliated to Firmicutes, order Clostridiales, family Caldicoprobacteraceae, with Caldicoprobacter oshimai (98.5%) being its closest relative. Based on phenotypic, phylogenetic, and genetic characteristics, strain TH7C1(T) is proposed as a novel species of genus Caldicoprobacter, Caldicoprobacter algeriensis, sp. nov. (strain TH7C1(T) = DSM 22661(T) = JCM 16184(T)).

  10. Caldicoprobacter guelmensis sp. nov., a thermophilic, anaerobic, xylanolytic bacterium isolated from a hot spring. (United States)

    Bouanane-Darenfed, Amel; Ben Hania, Wajdi; Hacene, Hocine; Cayol, Jean-Luc; Ollivier, Bernard; Fardeau, Marie-Laure


    A hyperthermophilic anaerobic bacterium, designated D2C22(T), was isolated from the hydrothermal hot spring of Guelma in north-east Algeria. The isolate was a Gram-stain-positive, non-sporulating, non-motile rod, appearing singly or in pairs (0.3-0.4 × 8.0-9.0 µm). Strain D2C22(T) grew anaerobically at 45-85 °C (optimum 65 °C), at pH 5-9 (optimum pH 6.8) and with 0-20 g NaCl l(-1). Strain D2C22(T) used glucose, galactose, lactose, fructose, ribose, xylose, arabinose, maltose, cellobiose, mannose, melibiose, sucrose, xylan and pyruvate (only in the presence of yeast extract or biotrypticase) as electron donors. The end products from glucose fermentation were acetate, lactate, CO2 and H2. Nitrate, nitrite, thiosulfate, elemental sulfur, sulfate and sulfite were not used as electron acceptors. The predominant cellular fatty acids were iso-C15:0 and iso-C17:0. The DNA G+C content was 41.6 mol%. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain D2C22(T) was most closely related to Caldicoprobacter oshimai JW/HY-331(T), Caldicoprobacter algeriensis TH7C1(T) and Acetomicrobium faecale DSM 20678(T) (95.5, 95.5 and 95.3% 16S rRNA gene sequence similarity, respectively). Based on phenotypic, phylogenetic and chemotaxonomic characteristics, strain D2C22(T) is proposed to be a representative of a novel species of the genus Caldicoprobacter within the order Clostridiales, for which the name Caldicoprobacter guelmensis sp. nov. is proposed. The type strain is D2C22(T) (=DSM 24605(T)=JCM 17646(T)).

  11. Vibrio japonicus sp. nov., a novel member of the Nereis clade in the genus Vibrio isolated from the coast of Japan. (United States)

    Doi, Hiroyasu; Osawa, Ikuko; Adachi, Hayamitsu; Kawada, Manabu


    A novel Vibrio strain, JCM 31412T, was isolated from seawater collected from the Inland Sea (Setonaikai), Japan, and characterized as a Gram-negative, oxidase-positive, catalase-negative, facultatively anaerobic, motile, ovoid-shaped bacterium with one polar flagellum. Based on 16S rDNA gene identity, strain JCM 31412T showed a close relationship with type strains of Vibrio brasiliensis (LMG 20546T, 98.2% identity), V. harveyi (NBRC 15634T, 98.2%), V. caribbeanicus (ATCC BAA-2122T, 97.8%) and V. proteolyticus (NBRC 13287T, 97.8%). The G+C content of strain JCM 31412T DNA was 46.8%. Multi-locus sequence analysis (MLSA) of eight loci (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA and topA; 5535bp) further clustered strain JCM 31412T in the Nereis clade, genus Vibrio. Phenotypically, strain JCM 31412T differed from the closest related Vibrio species in its utilization of melibiose and raffinose, and its lack of casein and gelatin hydrolysis. It was further differentiated based on its fatty acid composition, specifically properties of C12:03OH and summed features, which were significantly different from those of V. brasiliensis, V. nigripulchritudo and V. caribbeanicus type strains. Overall, the results of DNA-DNA hybridization, and physiological and biochemical analysis differentiated strain JCM 31412T from other described species of the genus Vibrio. Based on these polyphasic taxonomic findings, it was therefore concluded that JCM 31412T was a novel Vibrio species, for which the name Vibrio japonicus sp. nov. was proposed, with JCM 31412T (= LMG 29636T = ATCC TSD-62T) as the type strain.

  12. Binding profiles and cytokine-inducing effects of fish rhamnose-binding lectins on Burkitt's lymphoma Raji cells. (United States)

    Hosono, Masahiro; Sugawara, Shigeki; Matsuda, Atsushi; Tatsuta, Takeo; Koide, Yasuhiro; Hasan, Imtiaj; Hasan, Imtiaji; Ozeki, Yasuhiro; Nitta, Kazuo


    Rhamnose-binding lectin (RBL) is one of the animal lectin categories which take part in the innate immune responses of fish. Osmerus lanceolatus lectin (OLL) from shishamo smelt eggs is an RBL composed of two tandem-repeated domains, both of which are considered to be a carbohydrate-recognition domain. SAL, catfish (Silurus asotus) egg RBL composed of three domains, binds to Burkitt's lymphoma Raji cells through globotriaosylceramide (Gb3) carbohydrate chain and to reduce cell size and growth by altering membrane composition without causing cell death. In this experiment, we tried to compare the binding effects of these two RBLs on Raji cells. Flow cytometric and fluorescence microscopic analyses revealed that OLL also directly bound to and shrunk Raji cells with ten times less reactivity than SAL but reduced cell growth with decreasing cell viability. Anti-Gb3 antibody completely blocked the binding of SAL to Raji cells but not that of OLL. In addition, the direct bindings of OLL and SAL to Raji cells were comparably inhibited by melibiose, but lactose was more effective inhibitor for the binding of OLL than that of SAL. These results suggest that OLL has slightly different cell-binding property compared with SAL and binds not only to Gb3 but also to the other carbohydrate receptor-bearing β-galactoside chains. The quantitative RT-PCR analysis revealed that SAL induced the expression of TNF-α but not of IFN-γ, IL-1β, and IL-10. Thus, SAL-induced cytostatic effect on Raji cells might be partially caused by TNF-α-mediated signaling pathway.

  13. Vibrio japonicus sp. nov., a novel member of the Nereis clade in the genus Vibrio isolated from the coast of Japan.

    Directory of Open Access Journals (Sweden)

    Hiroyasu Doi

    Full Text Available A novel Vibrio strain, JCM 31412T, was isolated from seawater collected from the Inland Sea (Setonaikai, Japan, and characterized as a Gram-negative, oxidase-positive, catalase-negative, facultatively anaerobic, motile, ovoid-shaped bacterium with one polar flagellum. Based on 16S rDNA gene identity, strain JCM 31412T showed a close relationship with type strains of Vibrio brasiliensis (LMG 20546T, 98.2% identity, V. harveyi (NBRC 15634T, 98.2%, V. caribbeanicus (ATCC BAA-2122T, 97.8% and V. proteolyticus (NBRC 13287T, 97.8%. The G+C content of strain JCM 31412T DNA was 46.8%. Multi-locus sequence analysis (MLSA of eight loci (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA and topA; 5535bp further clustered strain JCM 31412T in the Nereis clade, genus Vibrio. Phenotypically, strain JCM 31412T differed from the closest related Vibrio species in its utilization of melibiose and raffinose, and its lack of casein and gelatin hydrolysis. It was further differentiated based on its fatty acid composition, specifically properties of C12:03OH and summed features, which were significantly different from those of V. brasiliensis, V. nigripulchritudo and V. caribbeanicus type strains. Overall, the results of DNA-DNA hybridization, and physiological and biochemical analysis differentiated strain JCM 31412T from other described species of the genus Vibrio. Based on these polyphasic taxonomic findings, it was therefore concluded that JCM 31412T was a novel Vibrio species, for which the name Vibrio japonicus sp. nov. was proposed, with JCM 31412T (= LMG 29636T = ATCC TSD-62T as the type strain.

  14. Bombella intestini gen. nov., sp. nov., an acetic acid bacterium isolated from bumble bee crop. (United States)

    Li, Leilei; Praet, Jessy; Borremans, Wim; Nunes, Olga C; Manaia, Célia M; Cleenwerck, Ilse; Meeus, Ivan; Smagghe, Guy; De Vuyst, Luc; Vandamme, Peter


    In the frame of a bumble bee gut microbiota study, acetic acid bacteria (AAB) were isolated using a combination of direct isolation methods and enrichment procedures. MALDI-TOF MS profiling of the isolates and a comparison of these profiles with profiles of established AAB species identified most isolates as Asaia astilbis or as 'Commensalibacter intestini', except for two isolates (R-52486 and LMG 28161(T)) that showed an identical profile. A nearly complete 16S rRNA gene sequence of strain LMG 28161(T) was determined and showed the highest pairwise similarity to Saccharibacter floricola S-877(T) (96.5%), which corresponded with genus level divergence in the family Acetobacteraceae. Isolate LMG 28161(T) was subjected to whole-genome shotgun sequencing; a 16S-23S rRNA internal transcribed spacer (ITS) sequence as well as partial sequences of the housekeeping genes dnaK, groEL and rpoB were extracted for phylogenetic analyses. The obtained data confirmed that this isolate is best classified into a new genus in the family Acetobacteraceae. The DNA G+C content of strain LMG 28161(T) was 54.9 mol%. The fatty acid compositions of isolates R-52486 and LMG 28161(T) were similar to those of established AAB species [with C18:1ω7c (43.1%) as the major component], but the amounts of fatty acids such as C19:0 cyclo ω8c, C14:0 and C14:0 2-OH enabled to differentiate them. The major ubiquinone was Q-10. Both isolates could also be differentiated from the known genera of AAB by means of biochemical characteristics, such as their inability to oxidize ethanol to acetic acid, negligible acid production from melibiose, and notable acid production from d-fructose, sucrose and d-mannitol. In addition, they produced 2-keto-d-gluconate, but not 5-keto-d-gluconate from d-glucose. Therefore, the name Bombella intestini gen nov., sp. nov. is proposed for this new taxon, with LMG 28161(T) ( =DSM 28636(T) =R-52487(T)) as the type strain of the type species. © 2015 IUMS.

  15. Rheological behaviour of some saccharides in aqueous potassium chloride solutions over temperature range (288.15 to 318.15) K

    Energy Technology Data Exchange (ETDEWEB)

    Banipal, Parampaul K., E-mail: [Department of Chemistry, Guru Nanak Dev University, Amritsar 143005 (India); Chahal, Amanpreet K.; Singh, Vickramjeet [Department of Chemistry, Guru Nanak Dev University, Amritsar 143005 (India); Banipal, Tarlok S. [Department of Applied Chemistry, Guru Nanak Dev University, Amritsar 143005 (India)


    The viscosities, {eta} of mono-, di-, tri-saccharides and methylglycosides, viz., D(+)-xylose (XYL), D(-)-arabinose (ARA), D(-)-ribose (RIB), D(-)-fructose (FRU), D(+)-galactose (GAL), D(+)-mannose (MAN), D(+)-glucose (GLU), D(+)-melibiose (MEL), D(+)-cellobiose (CEL), D(+)-lactose monohydrate (LAC), D(+)-maltose monohydrate (MAL), D(+)-trehalose dihydrate (TRE), sucrose (SUC), D(+)-raffinose pentahydrate (RAF), {alpha}-methyl-D(+)-glucoside ({alpha}-Me-GLU), methyl-{alpha}-D-xylopyranoside (Me-{alpha}-XYL), and methyl-{beta}-D-xylopyranoside (Me-{beta}-XYL) in water and in (0.5, 1.0, 2.0, and 3.0) mol . kg{sup -1} aqueous solutions of potassium chloride (KCl) have been determined at T = (288.15, 298.15, 308.15, and 318.15) K from efflux time measurements by using a capillary viscometer. Densities used to determine viscosities have been reported earlier. The viscosity data have been utilized to determine the viscosity B-coefficients employing the Jones-Dole equation at different temperatures. From these data, the viscosity B-coefficients of transfer, {Delta}{sub t}B have been estimated for the transfer of various saccharides/methylglycosides from water to aqueous potassium chloride solutions. The {Delta}{sub t}B values have been found to be positive, whose magnitude increases with the increase in concentration of potassium chloride in all cases. The dB/dT coefficients, pair, {eta}{sub AB} and triplet, {eta}{sub ABB} viscometric interaction coefficients have also been determined. Gibbs free energies of activation and related thermodynamic parameters of activation of viscous flow have been determined employing Feakin's transition-state theory. The signs and magnitudes of various parameters have been discussed in terms of solute-solute and solute-solvent interactions occurring in these solutions. The effect of substitution of -OH by methoxy group, -OCH{sub 3} has also been discussed.

  16. Effect of sodium acetate on the volumetric behaviour of some mono-, di-, and tri-saccharides in aqueous solutions over temperature range (288.15 to 318.15) K

    Energy Technology Data Exchange (ETDEWEB)

    Banipal, Parampaul K., E-mail: [Department of Chemistry, Guru Nanak Dev University, Amritsar 143 005 (India); Singh, Vickramjeet [Department of Chemistry, Guru Nanak Dev University, Amritsar 143 005 (India); Banipal, Tarlok S. [Department of Applied Chemistry, Guru Nanak Dev University, Amritsar 143 005 (India)


    The standard partial molar volumes, V{sub 2}{sup 0} at infinite dilution of eight monosaccharides [D(+)-xylose, D(-)-arabinose, D(-)-ribose, L(-)-sorbose, D(-)-fructose, D(+)-galactose, D(+)-glucose, and D(+)-mannose], six disaccharides [D(+)-cellobiose, sucrose, D(+)-melibiose, D(+)-lactose monohydrate, D(+)-trehalose dihydrate, and D(+)-maltose monohydrate] and two trisaccharides [D(+)-melizitose and D(+)-raffinose pentahydrate] (molalities of saccharides range from (0.03 to 0.12) mol . kg{sup -1}) have been determined in water and in (0.5, 1.0, 2.0, and 3.0) mol . kg{sup -1} aqueous sodium acetate solutions at temperatures, T = (288.15, 298.15, 308.15, and 318.15) K from density measurements using a vibrating-tube digital densimeter. From these results, corresponding standard partial molar volumes of transfer, DELTA{sub t}V{sub 2}{sup 0} have been determined for the transfer of various saccharides from water to aqueous solutions of sodium acetate. Positive values of DELTA{sub t}V{sub 2}{sup 0} were obtained for most of the saccharides, whose magnitude increase with the concentration of sodium acetate as well as temperature. However, negative DELTA{sub t}V{sub 2}{sup 0} values were observed for L(-)-sorbose, D(-)-fructose and D(+)-xylose at lower concentrations of co-solute. The negative magnitude of DELTA{sub t}V{sub 2}{sup 0} values decrease with rise of temperature from (288.15 to 318.15) K. Pair and higher order volumetric interaction coefficients have been determined by using McMillan-Mayer theory. Partial molar expansion coefficients, (partial derivV{sub 2}{sup 0}/partial derivT){sub p} and the second derivatives (partial deriv{sup 2}V{sub 2}{sup 0}/partial derivT{sup 2}){sub p} have also been estimated. These parameters have been utilized to understand various mixing effects in aqueous solutions due to the interactions between solute (saccharide) and co-solute (sodium acetate).

  17. Vibrio communis sp. nov., isolated from the marine animals Mussismilia hispida, Phyllogorgia dilatata, Palythoa caribaeorum, Palythoa variabilis and Litopenaeus vannamei. (United States)

    Chimetto, Luciane A; Cleenwerck, Ilse; Alves, Nelson; Silva, Bruno Sergio; Brocchi, Marcelo; Willems, Anne; De Vos, Paul; Thompson, Fabiano L


    Eight Vibrio isolates originating from the marine corals Mussismilia hispida and Phyllogorgia dilatata and the zoanthids Palythoa caribaeorum and Palythoa variabilis in Brazil and the Pacific white shrimp (Litopenaeus vannamei) in Ecuador were studied by means of a polyphasic approach. The novel isolates formed a tight monophyletic group in the genus Vibrio and were closely related to species of the Vibrio harveyi group, to which they showed more than 99 % 16S rRNA gene sequence similarity. Analysis based on concatenated sequences of the following seven genes, 16S rRNA, gyrB, recA, rpoA, topA, pyrH and mreB (5633 bp in length), showed clear separation between the isolates and species of the V. harveyi group. Amplified fragment length polymorphism (AFLP) analysis, performed previously, revealed that a representative isolate of this group, LMG 20370, was clearly separate from known Vibrio species (it belonged to the so-called AFLP cluster A31). DNA-DNA hybridization (DDH) experiments with representative isolates and type strains of the V. harveyi species group revealed high DDH between the novel isolates (more than 74 %) and less than 70 % DDH towards type strains of related Vibrio species, proving the novel species status of the isolates. Phenotypically, the novel species belongs to the arginine dihydrolase (A)-negative, lysine decarboxylase (L)-positive and ornithine decarboxylase (O)-positive (A-/L+/O+) cluster reported previously. Most species of the V. harveyi group (i.e. Vibrio rotiferianus, V. harveyi, V. parahaemolyticus and V. alginolyticus) also belong to this A-/L+/O+ cluster. However, several phenotypic features can be used for the identification of the novel species. In contrast to its closest phylogenetic neighbours, the novel species exhibits esterase (C4) and N-acetyl-β-glucosaminidase activities, but it does not produce acetoin, does not use citrate, α-ketoglutaric acid or propionic acid and does not ferment melibiose. The novel species can

  18. Enterobacter asburiae sp. nov., a new species found in clinical specimens, and reassignment of Erwinia dissolvens and Erwinia nimipressuralis to the genus Enterobacter as Enterobacter dissolvens comb. nov. and Enterobacter nimipressuralis comb. nov. (United States)

    Brenner, D J; McWhorter, A C; Kai, A; Steigerwalt, A G; Farmer, J J


    Enterobacter asburiae sp. nov. is a new species that was formerly referred to as Enteric Group 17 and that consists of 71 strains, 70 of which were isolated from humans. Enterobacter asburiae sp. nov. strains gave positive reactions in tests for methyl red, citrate utilization (Simmons and Christensen's), urea hydrolysis, L-ornithine decarboxylase, growth in KCN, acid and gas production from D-glucose, and acid production from L-arabinose, cellobiose, glycerol (negative in 1 to 2 days, positive in 3 to 7 days), lactose, D-mannitol, alpha-methyl-D-glucoside, salicin, D-sorbitol, sucrose, trehalose, and D-xylose. They gave negative reactions in the Voges-Proskauer test and in tests for indole, H2S production, phenylalanine, L-lysine decarboxylase, motility, gelatin, utilization of malonate, lipase, DNase, tyrosine clearing, acid production from adonitol, D-arabitol, dulcitol, erythritol, i(myo)-inositol, melibiose, and L-rhamnose. They gave variable reactions in tests for L-arginine dihydrolase (25% positive after 2 days) and acid production from raffinose (69% positive after 2 days). Thirty-four Enterobacter asburiae sp. nov. strains were tested for DNA relatedness by the hydroxyapatite method with 32PO4-labeled DNA from the designated type strain (1497-78, ATCC 35953). The strains were 69 to 100% related in 60 degrees C reactions and 63 to 100% related in 75 degrees C reactions. Divergence within related sequences was 0 to 2.5%. Relatedness of Enterobacter asburiae sp. nov. to 84 strains of members of the Enterobacteriaceae was 5 to 63%, with closest relatedness to strains of Enterobacter cloacae, Erwinia dissolvens, Enterobacter taylorae, Enterobacter agglomerans, Erwinia nimipressuralis, and Enterobacter gergoviae. All strains tested were susceptible to gentamicin and sulfdiazine, and most were susceptible to chloramphenicol, colistin, kanamycin, nalidixic acid, carbenicillin and streptomycin. All strains were resistant to ampicillan, cephalothin, and penicillin

  19. Studies on volumetric properties of some saccharides in aqueous potassium chloride solutions over temperature range (288.15 to 318.15) K

    Energy Technology Data Exchange (ETDEWEB)

    Banipal, Parampaul K. [Department of Chemistry, Guru Nanak Dev University, Amritsar 143 005 (India)], E-mail:; Chahal, Amanpreet K. [Department of Chemistry, Guru Nanak Dev University, Amritsar 143 005 (India); Banipal, Tarlok S. [Department of Applied Chemistry, Guru Nanak Dev University, Amritsar 143 005 (India)


    The standard partial molar volumes, V{sub 2}{sup {infinity}} at infinite dilution of monosaccharides; D(+)-xylose, D(-)-arabinose, D(-)-ribose, D(+)-mannose, D(+)-galactose, D(-)-fructose and D(+)-glucose, disaccharides; D(+)-melibiose, D(+)-cellobiose, D(+)-maltose monohydrate, D(+)-trehalose dihydrate, D(+)-lactose monohydrate and sucrose, trisaccharide; D(+)-raffinose pentahydrate, methylglycosides; {alpha}-methyl-D(+)-glucoside, methyl-{alpha}-D-xylopyranoside and methyl-{beta}-D-xylopyranoside have been determined in water and in aqueous solutions of potassium chloride (0.5, 1.0, 2.0, and 3.0) mol . kg{sup -1} at T = (288.15, 298.15, 308.15, and 318.15) K from density measurements employing a vibrating-tube densimeter. These results have been utilized to determine the corresponding standard partial molar volumes of transfer, {delta}{sub t}V{sub 2}{sup {infinity}} for the transfer of various saccharides from water to aqueous potassium chloride solutions. The standard transfer volumes have been found to be positive (except for {alpha}- and {beta}-methyl xylopyranosides in 0.5 mol . kg{sup -1} solutions of potassium chloride) whose magnitude increase with the concentration of potassium chloride as well as temperature for all the saccharides. Partial molar expansion coefficients, ({partial_derivative}V{sub 2}{sup {infinity}}/{partial_derivative}T){sub p} and the second derivative ({partial_derivative}{sup 2}V{sub 2}{sup {infinity}}/{partial_derivative}T{sup 2}){sub p} values have been estimated. Pair and higher order volumetric interaction coefficients have also been calculated from {delta}{sub t}V{sub 2}{sup {infinity}} by using the McMillan-Mayer theory. These parameters have been discussed in terms of the solute-cosolute interactions and are used to understand various mixing effects due to these interactions. The effect of substitution of -OH by glycosidic group, -OCH{sub 3} is also discussed. Attempt has also been made to discuss the stereochemical effects

  20. A chromogenic plating medium for the isolation and identification of Enterobacter sakazakii from foods, food ingredients, and environmental sources. (United States)

    Restaino, L; Frampton, E W; Lionberg, W C; Becker, R J


    A chromogenic agar, R&F Enterobacter sakazakii chromogenic plating medium (ESPM), was developed for isolating presumptive colonies of E. sakazakii from foods and environmental sources. ESPM contains two chromogenic substrates (5-bromo-4-chloro-3-indoxyl-alpha-D-glucopyranoside and 5-bromo-4-chloro-3-indoxyl-beta-D-cellobioside), three sugars (sorbitol, D-arabitol, and adonitol), a pH indicator, and inhibitors (bile salts, vancomycin, and cefsulodin), which all contribute to its selectivity and differential properties. On ESPM, 79 pure culture strains of E. sakazakii (10 clinical isolates and others from food and environmental sources) yielded blue-black (three strains were blue-gray) raised colonies, 1 to 2 mm in diameter with and without halos after 24 h at 35 degrees C. Other enteric organisms plus Pseudomonas aeruginosa yielded white, yellow, green, or clear colonies with and without clear halos. Of these genera, only Shigella sonnei and one Pantoea strain produced blue-black to blue-gray colonies. ESPM was used to isolate E. sakazakii from a variety of foods: corn, wheat, and rice flours; powdered infant formula; dairy products (dried milk, whey, and caseinates); cereals; and environmental sources. Most false-positive results on ESPM were eliminated by observing acid production on either sucrose or melibiose after 6 h at 35 degrees C on a R&F E. sakazakii screening medium (ESSM) biplate. In an analysis of 240 samples, the number of samples positive for E. sakazakii by the ESPM-ESSM method and the U.S. Food and Drug Administration protocols (violet red bile glucose agar and tryptic soy agar) were 27 and 16, respectively, with sensitivity and specificity values of 100.0 and 96.9% versus 59.3 and 43.7%, respectively. These data support the fact that E. sakazakii confirmation should be based on more than one confirmation system. Both the API 20E and Biolog Microlog3 4.20 systems should be used for confirmation of E. sakazakii isolates.

  1. Improving Cotton Embryo Culture by Simulating In Ovulo Nutrient and Hormone Levels

    Energy Technology Data Exchange (ETDEWEB)

    Rodney Fuller; Vincent Liddiard; J. Hess; John Carman


    Plant ovules provide zygotes with a physicochemical environment that supports embryo differentiation, growth, and maturation. The exact nature of this embryogenesis-enabling environment is not well characterized, as evidenced by failed attempts to induce normal embryony from zygotes or proembryos (precotyledonary) on defined media. To identify factors required for cotton (Gossypium hirsutum L.) zygotic embryony in vitro, we previously performed chemical and dissolved oxygen tension analyses of cotton ovule fluids and tissues at multiple stages of embryony in situ. Based on these analyses, we report herein the development of procedures that normalize embryo differentiation, growth, maturation, and germination in vitro, starting with proembryos. Our medium differed from Murashige and Skoog (MS) medium as follows (percentage of MS): N (30%, mostly from ten amino acids), P (815%), K (237%), Mg (85%), Ca (267%), S (506%), Fe (88%), and myoinositol (883%). Levels of other MS nutrients and vitamins, except sucrose, were kept at MS levels. Additionally, we included 100 mg L-1 casein hydrolysate plus the following (mmol L-1): d-glucose (1.8), fructose (4.7), sucrose (62.0), arabinose (7.1), melibiose (3.5), malic acid (11.6), and citric acid (3.8). Mannitol was added to achieve a medium osmotic potential of -1.10 MPa, and an atmospheric O2 tension of 3.3 mol m-3 at the surface of embryos was maintained during culture. When cultured on medium containing 8.0 µmol L-1 indole-3-acetic acid, 80-90% of proembryos (as small as 100 cells) of cultivars HS-26 and B-27 increased four- to eightfold in surface area during the first 18 d in culture and germinated thereafter to produce viable plants. Increases in surface area of proembryos cultured on a modified MS medium previously used for somatic embryogenesis were from 0.2- to 0.6-fold. The described embryo culture medium should be useful for studying nutritional and molecular aspects of early embryony and possibly for plant zygote