Method for Making High Molecular Weight, Extended pi-Conjugated Polymers

DTIC Science & Technology

2001-05-04

derivatized poly(terephthalates)s as coatings for electronics components, and as construction materials for field- effect transistors, both applications...mannose, dulose, idose, galactose and talose; ketoses such as erythrulose, ribulose, xylulose, psicose, fructose, sorbose, tagatose ; di-, tri-, 12...show that Sc(OTr)3 alone was not effective as a 15 polymerization catalyst. When the organic salt was introduced, Sc(OTr)3 became marginally

[Crabtree effect caused by ketoses in isolated rat hepatocytes].

PubMed

Martínez, P; Carrascosa, J M; Núñez de Castro, I

1982-01-01

Oxygen uptake and glycolytic activity were studied in hepatocytes isolated from fed rats. The addition of fructose or tagatose resulted in a 38% and 31% inhibition of cellular respiration respectively. The addition of 10 mM D-glyceraldehyde caused a slight Crabtree effect. Glucose, L-sorbose, or glycerol failed to modify oxygen consumption. Only incubation in the presence of fructose showed a high aerobic glycolysis measured by lactate production.

40 CFR 711.6 - Chemical substances for which information is not required.

Code of Federal Regulations, 2013 CFR

2013-07-01

... Partially Exempt Chemical Substances CASRN Chemical 50-70-4 D-glucitol. 50-81-7 L-ascorbic acid. 50-99-7 D-glucose. 56-81-5 1,2,3-Propanetriol. 56-87-1 L-lysine. 57-50-1 .alpha.-D-Glucopyranoside, .beta.-D...- trimethyltridecyl]-, (2R)-. 59-51-8 Methionine. 69-65-8 D-mannitol. 87-79-6 L-sorbose. 87-99-0 Xylitol. 96-10-6...

40 CFR 711.6 - Chemical substances for which information is not required.

Code of Federal Regulations, 2012 CFR

2012-07-01

... Partially Exempt Chemical Substances CASRN Chemical 50-70-4 D-glucitol. 50-81-7 L-ascorbic acid. 50-99-7 D-glucose. 56-81-5 1,2,3-Propanetriol. 56-87-1 L-lysine. 57-50-1 .alpha.-D-Glucopyranoside, .beta.-D...- trimethyltridecyl]-, (2R)-. 59-51-8 Methionine. 69-65-8 D-mannitol. 87-79-6 L-sorbose. 87-99-0 Xylitol. 96-10-6...

40 CFR 711.6 - Chemical substances for which information is not required.

Code of Federal Regulations, 2014 CFR

2014-07-01

... Partially Exempt Chemical Substances CASRN Chemical 50-70-4 D-glucitol. 50-81-7 L-ascorbic acid. 50-99-7 D-glucose. 56-81-5 1,2,3-Propanetriol. 56-87-1 L-lysine. 57-50-1 .alpha.-D-Glucopyranoside, .beta.-D...- trimethyltridecyl]-, (2R)-. 59-51-8 Methionine. 69-65-8 D-mannitol. 87-79-6 L-sorbose. 87-99-0 Xylitol. 96-10-6...

Metabolic engineering pathways for rare sugars biosynthesis, physiological functionalities, and applications-a review.

PubMed

Bilal, Muhammad; Iqbal, Hafiz M N; Hu, Hongbo; Wang, Wei; Zhang, Xuehong

2017-06-29

Biomolecules like rare sugars and their derivatives are referred to as monosaccharides particularly uncommon in nature. Remarkably, many of them have various known physiological functions and biotechnological applications in cosmetics, nutrition, and pharmaceutical industries. Also, they can be exploited as starting materials for synthesizing fascinating natural bioproducts with significant biological activities. Regrettably, most of the rare sugars are quite expensive, and their synthetic chemical routes are both limited and economically unfeasible due to expensive raw materials. On the other hand, their production by enzymatic means often suffers from low space-time yields and high catalyst costs due to hasty enzyme denaturation/degradation. In this context, biosynthesis of rare sugars with industrial importance is receiving renowned scientific attention, across the globe. Moreover, the utilization of renewable resources as energy sources via microbial fermentation or microbial metabolic engineering has appeared a new tool. This article presents a comprehensive review of physiological functions and biotechnological applications of rare ketohexoses and aldohexoses, including D-psicose, D-tagatose, L-tagatose, D-sorbose, L-fructose, D-allose, L-glucose, D-gulose, L-talose, L-galactose, and L-fucose. Novel in-vivo recombination pathways based on aldolase and phosphatase for the biosynthesis of rare sugars, particularly D-psicose and D-sorbose using robust microbial strains are also deliberated.

Synthesis of rare sugars with L-fuculose-1-phosphate aldolase (FucA) from Thermus thermophilus HB8.

PubMed

Li, Zijie; Cai, Li; Qi, Qingsheng; Styslinger, Thomas J; Zhao, Guohui; Wang, Peng George

2011-09-01

We report herein a one-pot four-enzyme approach for the synthesis of the rare sugars d-psicose, d-sorbose, l-tagatose, and l-fructose with aldolase FucA from a thermophilic source (Thermus thermophilus HB8). Importantly, the cheap starting material DL-GP (DL-glycerol 3-phosphate), was used to significantly reduce the synthetic cost. Copyright © 2011 Elsevier Ltd. All rights reserved.

Stepwise metabolic engineering of Gluconobacter oxydans WSH-003 for the direct production of 2-keto-L-gulonic acid from D-sorbitol.

PubMed

Gao, Lili; Hu, Yudong; Liu, Jie; Du, Guocheng; Zhou, Jingwen; Chen, Jian

2014-07-01

2-Keto-L-gulonic acid (2-KLG), the direct precursor of vitamin C, is currently produced by a two-step fermentation route from D-sorbitol. However, this route involves three bacteria, making the mix-culture system complicated and redundant. Thus, replacement of the conventional two-step fermentation process with a one-step process could be revolutionary in vitamin C industry. In this study, different combinations of five L-sorbose dehydrogenases (SDH) and two L-sorbosone dehydrogenases (SNDH) from Ketogulonicigenium vulgare WSH-001 were introduced into Gluconobacter oxydans WSH-003, an industrial strain used for the conversion of d-sorbitol to L-sorbose. The optimum combination produced 4.9g/L of 2-KLG. In addition, 10 different linker peptides were used for the fusion expression of SDH and SNDH in G. oxydans. The best recombinant strain (G. oxydans/pGUC-k0203-GS-k0095) produced 32.4g/L of 2-KLG after 168h. Furthermore, biosynthesis of pyrroloquinoline quinine (PQQ), a cofactor of those dehydrogenases, was enhanced to improve 2-KLG production. With the stepwise metabolic engineering of G. oxydans, the final 2-KLG production was improved to 39.2g/L, which was 8.0-fold higher than that obtained using independent expression of the dehydrogenases. These results bring us closer to the final one-step industrial-scale production of vitamin C. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Accumulation of 2-Keto-L-Gulonate at 33°C by a Thermotolerant Gluconobacter Oxydans Mutant Obtained by Ion Beam Implantation

NASA Astrophysics Data System (ADS)

Yan, Bing; Xu, An; Zhang, Wan; Zhou, Wei; Wang, Jun; Yao, Jianming; Yu, Zengliang

2006-03-01

To obtain thermotolerant mutants of G. oxydans, which can enhance the transformation rate of L-sorbose to 2-Keto-L-gulonate (2-KLG) at 33oC in a two-step process of vitamin C manufacture, ion beam was used as a mutation source. Gluconobacter oxydans G0 and Bacillus megaterium B0 were used in this study. The original strain Gluconobacter oxydans G0 was mutated by the heavy ion implantation facility at the Institute of Plasma Physics, Chinese Academy of Sciences. Several mutants including Gluconobacter oxydans GI13 were isolated and cocultured with Bacillus megaterium B0 at 33oC in shaking flasks. The average transformation rate of the new mixed strain GI13-B0 in per gram-molecule reached 94.4% after seven passages in shaking flasks, which was increased by 7% when compared with the original mixed strain G0-B0 (Gluconobacter oxydans G0 and Bacillus megaterium B0). Moreover, the transformation rate of I13B0 was stable at 94% at temperatures ranging from 25oC to 33oC, which would be of much value in reducing energy consumption in the manufacture of L-ascorbic acid, especially in the season of summer. To clarify some mechanism of the mutation, the specific activities of L-sorbose dehydrogenase in both G0 and GI13 were estimated.

Potential anthelmintic: D-psicose inhibits motility, growth and reproductive maturity of L1 larvae of Caenorhabditis elegans.

PubMed

Sato, Masashi; Kurose, Hiroyuki; Yamasaki, Toru; Izumori, Ken

2008-04-01

No anthelmintic sugars have yet been identified. Eight ketohexose stereoisomers (D- and L-forms of psicose, fructose, tagatose and sorbose), along with D-galactose and D-glucose, were examined for potency against L1 stage Caenorhabditis elegans fed Escherichia coli. Of the sugars, D-psicose specifically inhibited the motility, growth and reproductive maturity of the L1 stage. D-Psicose probably interferes with the nematode nutrition. The present results suggest that D-psicose, one of the rare sugars, is a potential anthelmintic.

Biosynthesis of rare ketoses through constructing a recombination pathway in an engineered Corynebacterium glutamicum.

PubMed

Yang, Jiangang; Zhu, Yueming; Li, Jitao; Men, Yan; Sun, Yuanxia; Ma, Yanhe

2015-01-01

Rare sugars have various known biological functions and potential for applications in pharmaceutical, cosmetics, and food industries. Here we designed and constructed a recombination pathway in Corynebacterium glutamicum, in which dihydroxyacetone phosphate (DHAP), an intermediate of the glycolytic pathway, and a variety of aldehydes were condensed to synthesize rare ketoses sequentially by rhamnulose-1-phosphate aldolase (RhaD) and fructose-1-phosphatase (YqaB) obtained from Escherichia coli. A wild-type strain harboring this artificial pathway had the ability to produce D-sorbose and D-psicose using D-glyceraldehyde and glucose as the substrates. The tpi gene, encoding triose phosphate isomerase was further deleted, and the concentration of DHAP increased to nearly 20-fold relative to that of the wild-type. After additional optimization of expression levels from rhaD and yqaB genes and of the fermentation conditions, the engineered strain SY6(pVRTY) exhibited preferable performance for rare ketoses production. Its yield increased to 0.59 mol/mol D-glyceraldehyde from 0.33 mol/mol D-glyceraldehyde and productivity to 2.35 g/L h from 0.58 g/L h. Moreover, this strain accumulated 19.5 g/L of D-sorbose and 13.4 g/L of D-psicose using a fed-batch culture mode under the optimal conditions. In addition, it was verified that the strain SY6(pVRTY) meanwhile had the ability to synthesize C4, C5, C6, and C7 rare ketoses when a range of representative achiral and homochiral aldehydes were applied as the substrates. Therefore, the platform strain exhibited the potential for microbial production of rare ketoses and deoxysugars. © 2014 Wiley Periodicals, Inc.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sanctis, Daniele de; Rêgo, Ana T.; Marçal, David

The sorbitol operon regulator from K. pneumoniae has been overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 3.2 Å. The sorbitol operon regulator (SorC) regulates the metabolism of l-sorbose in Klebsiella pneumonia. SorC was overexpressed in Escherichia coli and purified, and crystals were obtained of a tetrameric form. A single crystal showed X-ray diffraction to 3.20 Å. The crystal belongs to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 91.6, b = 113.3, c = 184.1 Å. Analysis of the molecular-replacement solution indicates the presence of four SorC molecules in the asymmetric unit.

Sweet Structural Signatures Unveiled in Ketohexoses.

PubMed

Bermúdez, Celina; Peña, Isabel; Mata, Santiago; Alonso, José L

2016-11-14

The conformational behaviour of naturally occurring ketohexoses has been revealed in a supersonic expansion by Fourier transform microwave spectroscopy coupled with a laser ablation source. Three, two and one conformers of d-tagatose, d-psicose and l-sorbose, respectively, have been identified by their rotational constants extracted from the analysis of the spectra. Singular structural signatures involving the hydroxyl groups OH (1) and OH (2) have been disentangled from the intricate intramolecular hydrogen bond networks stabilising the most abundant conformers. The present results place the old Shallenberger and Kier sweetness theories on a firmer footing. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Vibrational Raman optical activity of ketose monosaccharides

NASA Astrophysics Data System (ADS)

Bell, Alasdair F.; Hecht, Lutz; Barron, Laurence D.

1995-07-01

The vibrational Raman optical activity (ROA) spectra of the four ketose sugars D-fructose, L-sorbose, D-tagatose and D-psicose in aqueous solution, which have been measured in backscattering in the range ≈250-1500 cm -1, are reported. These results are combined with those from a previous ROA study of aldose and pentose sugars in an attempt to establish new vibrational assignments and to verify old ones. The high information content of these spectra provides a new perspective on all the central features of monosaccharide stereochemistry including dominant anomeric configuration, ring conformation, exocyclic CH 2OH group conformation and relative disposition of the hydroxyl groups around the ring.

Unveiling the Sweet Conformations of Ketohexoses

NASA Astrophysics Data System (ADS)

Bermudez, C.; Pena, I.; Cabezas, C.; Daly, A. M.; Mata, S.; Alonso, J. L.

2013-06-01

The conformational behavior of ketohexoses D-Fructose, L-Sorbose, D-Tagatose and D-Psicose has been revealed from their rotational spectra. A broadband microwave spectrometer (CP-FTMW) has been used to rapidly acquire the rotational spectra in the 6 to 12 GHz frequency range. All observed species are stabilized by complicated intramolecular hydrogen-bonding networks. Structural motifs related to the sweetness of ketohexoses are revealed. G. G. Brown, B. C. Dian, K. O. Douglass, S. M. Geyer, S. T. Shipman, B. H. Pate, Rev. Sci. Instrum. 2008, 79, 053103. S. Mata, I. Peña, C. Cabezas, J. C. López, J. L. Alonso, J. Mol. Spectrosc. 2012, 280, 91.

Characterization of D-tagatose-3-epimerase from Rhodobacter sphaeroides that converts D-fructose into D-psicose.

PubMed

Zhang, Longtao; Mu, Wanmeng; Jiang, Bo; Zhang, Tao

2009-06-01

A non-characterized gene, previously proposed as the D-tagatose-3-epimerase gene from Rhodobacter sphaeroides, was cloned and expressed in Escherichia coli. Its molecular mass was estimated to be 64 kDa with two identical subunits. The enzyme specificity was highest with D-fructose and decreased for other substrates in the order: D-tagatose, D-psicose, D-ribulose, D-xylulose and D-sorbose. Its activity was maximal at pH 9 and 40 degrees C while being enhanced by Mn(2+). At pH 9 and 40 degrees C, 118 g D-psicose l(-1) was produced from 700 g D-fructose l(-1) after 3 h.

Enzymatic synthesis of rare sugars with L-rhamnulose-1-phosphate aldolase from Thermotoga maritima MSB8.

PubMed

Li, Zijie; Wu, Xiaoru; Cai, Li; Duan, Shenglin; Liu, Jia; Yuan, Peng; Nakanishi, Hideki; Gao, Xiao-Dong

2015-09-15

L-Rhamnulose-1-phosphate aldolase from a thermophilic source (Thermotoga maritima MSB8) (RhaDT.mari) was heterologously overexpressed in Escherichia coli and the stereoselectivity of this enzyme with or without Nus tag was investigated. We also applied this enzyme to the synthesis of rare sugars D-psicose, D-sorbose, L-tagatose and L-fructose using our one-pot four-enzyme system. To the best of our knowledge, this is the first use of RhaD from a thermophilic source for rare sugar synthesis and the temperature tolerance of this enzyme paves the path for large scale fermentation. Copyright © 2015 Elsevier Ltd. All rights reserved.

Correlation of Lactobacillus rhamnosus Genotypes and Carbohydrate Utilization Signatures Determined by Phenotype Profiling

PubMed Central

Lambert, Jolanda; van Limpt, Kees; Wels, Michiel; Smokvina, Tamara; Knol, Jan; Kleerebezem, Michiel

2015-01-01

Lactobacillus rhamnosus is a bacterial species commonly colonizing the gastrointestinal (GI) tract of humans and also frequently used in food products. While some strains have been studied extensively, physiological variability among isolates of the species found in healthy humans or their diet is largely unexplored. The aim of this study was to characterize the diversity of carbohydrate utilization capabilities of human isolates and food-derived strains of L. rhamnosus in relation to their niche of isolation and genotype. We investigated the genotypic and phenotypic diversity of 25 out of 65 L. rhamnosus strains from various niches, mainly human feces and fermented dairy products. Genetic fingerprinting of the strains by amplified fragment length polymorphism (AFLP) identified 11 distinct subgroups at 70% similarity and suggested niche enrichment within particular genetic clades. High-resolution carbohydrate utilization profiling (OmniLog) identified 14 carbon sources that could be used by all of the strains tested for growth, while the utilization of 58 carbon sources differed significantly between strains, enabling the stratification of L. rhamnosus strains into three metabolic clusters that partially correlate with the genotypic clades but appear uncorrelated with the strain's origin of isolation. Draft genome sequences of 8 strains were generated and employed in a gene-trait matching (GTM) analysis together with the publicly available genomes of L. rhamnosus GG (ATCC 53103) and HN001 for several carbohydrates that were distinct for the different metabolic clusters: l-rhamnose, cellobiose, l-sorbose, and α-methyl-d-glucoside. From the analysis, candidate genes were identified that correlate with l-sorbose and α-methyl-d-glucoside utilization, and the proposed function of these genes could be confirmed by heterologous expression in a strain lacking the genes. This study expands our insight into the phenotypic and genotypic diversity of the species L. rhamnosus and explores the relationships between specific carbohydrate utilization capacities and genotype and/or niche adaptation of this species. PMID:26048937

Complete Hexose Isomer Identification with Mass Spectrometry

NASA Astrophysics Data System (ADS)

Nagy, Gabe; Pohl, Nicola L. B.

2015-04-01

The first analytical method is presented for the identification and absolute configuration determination of all 24 aldohexose and 2-ketohexose isomers, including the D and L enantiomers for allose, altrose, galactose, glucose, gulose, idose, mannose, talose, fructose, psicose, sorbose, and tagatose. Two unique fixed ligand kinetic method combinations were discovered to create significant enough energetic differences to achieve chiral discrimination among all 24 hexoses. Each of these 24 hexoses yields unique ratios of a specific pair of fragment ions that allows for simultaneous determination of identification and absolute configuration. This mass spectrometric-based methodology can be readily employed for accurate identification of any isolated monosaccharide from an unknown biological source. This work provides a key step towards the goal of complete de novo carbohydrate analysis.

Glycation of myofibrillar proteins and ATPase activity after incubation with eleven sugars.

PubMed

Syrový, I

1994-01-01

Rat skeletal muscle myofibrils were incubated in the presence of D-glucose, D-fructose, D-galactose, D-ribose, D-tagatose, D-arabinose, D-xylose, D-mannose, L-sorbose, L-rhamnose or DL-glyceraldehyde and myofibrillar ATPase activity as well as the extent of glycation was measured. The attachment of sugars to proteins during glycation was generally dependent on the percentage of a given sugar present in the open-chain form. Glycation resulted in the decrease of myofibrillar ATPase activity. This decrease was low after incubation of myofibrillar proteins with slowly glycating sugars (e.g. glucose) and high with fast glycating sugars (e.g. ribose or glyceraldehyde). ATPase activity was less reduced in the presence of beta-mercaptoethanol.

Biosynthesis of rare hexoses using microorganisms and related enzymes

PubMed Central

Li, Zijie; Gao, Yahui; Nakanishi, Hideki

2013-01-01

Summary Rare sugars, referred to as monosaccharides and their derivatives that rarely exist in nature, can be applied in many areas ranging from foodstuffs to pharmaceutical and nutrition industry, or as starting materials for various natural products and drug candidates. Unfortunately, an important factor restricting the utilization of rare sugars is their limited availability, resulting from limited synthetic methods. Nowadays, microbial and enzymatic transformations have become a very powerful tool in this field. This article reviews the biosynthesis and enzymatic production of rare ketohexoses, aldohexoses and sugar alcohols (hexitols), including D-tagatose, D-psicose, D-sorbose, L-tagatose, L-fructose, 1-deoxy-L-fructose, D-allose, L-glucose, L-talose, D-gulose, L-galactose, L-fucose, allitol, D-talitol, and L-sorbitol. New systems and robust catalysts resulting from advancements in genomics and bioengineering are also discussed. PMID:24367410

Biosynthesis of rare hexoses using microorganisms and related enzymes.

PubMed

Li, Zijie; Gao, Yahui; Nakanishi, Hideki; Gao, Xiaodong; Cai, Li

2013-11-12

Rare sugars, referred to as monosaccharides and their derivatives that rarely exist in nature, can be applied in many areas ranging from foodstuffs to pharmaceutical and nutrition industry, or as starting materials for various natural products and drug candidates. Unfortunately, an important factor restricting the utilization of rare sugars is their limited availability, resulting from limited synthetic methods. Nowadays, microbial and enzymatic transformations have become a very powerful tool in this field. This article reviews the biosynthesis and enzymatic production of rare ketohexoses, aldohexoses and sugar alcohols (hexitols), including D-tagatose, D-psicose, D-sorbose, L-tagatose, L-fructose, 1-deoxy-L-fructose, D-allose, L-glucose, L-talose, D-gulose, L-galactose, L-fucose, allitol, D-talitol, and L-sorbitol. New systems and robust catalysts resulting from advancements in genomics and bioengineering are also discussed.

A rare sugar, d-allose, confers resistance to rice bacterial blight with upregulation of defense-related genes in Oryza sativa.

PubMed

Kano, Akihito; Gomi, Kenji; Yamasaki-Kokudo, Yumiko; Satoh, Masaru; Fukumoto, Takeshi; Ohtani, Kouhei; Tajima, Shigeyuki; Izumori, Ken; Tanaka, Keiji; Ishida, Yutaka; Tada, Yasuomi; Nishizawa, Yoko; Akimitsu, Kazuya

2010-01-01

We investigated responses of rice plant to three rare sugars, d-altrose, d-sorbose, and d-allose, due to establishment of mass production methods for these rare sugars. Root growth and shoot growth were significantly inhibited by d-allose but not by the other rare sugars. A large-scale gene expression analysis using a rice microarray revealed that d-allose treatment causes a high upregulation of many defense-related, pathogenesis-related (PR) protein genes in rice. The PR protein genes were not upregulated by other rare sugars. Furthermore, d-allose treatment of rice plants conferred limited resistance of the rice against the pathogen Xanthomonas oryzae pv. oryzae but the other tested sugars did not. These results indicate that d-allose has a growth inhibitory effect but might prove to be a candidate elicitor for reducing disease development in rice.

Effects of topical application of aqueous solutions of hexoses on epidermal permeability barrier recovery rate after barrier disruption.

PubMed

Denda, Mitsuhiro

2011-11-01

Previous studies have suggested that hexose molecules influence the stability of phospholipid bilayers. Therefore, the effects of topical application of all 12 stereoisomers of dextro-hexose on the epidermal barrier recovery rate after barrier disruption were evaluated. Immediately after tape stripping, 0.1 m aqueous solution of each hexose was applied on hairless mouse skin. Among the eight dextro-aldohexoses, topical application of altose, idose, mannose and talose accelerated the barrier recovery, while allose, galactose, glucose and gulose had no effect. Among the four dextro-ketohexoses, psicose, fructose, sorbose and tagatose all accelerated the barrier recovery. As the effects of hexoses on the barrier recovery rate appeared within 1 h, the mechanism is unlikely to be genomic. Instead, these hexoses may influence phase transition of the lipid bilayers of lamellar bodies and cell membrane, a crucial step in epidermal permeability barrier homeostasis. © 2011 John Wiley & Sons A/S.

[Kinetic properties of the fructose influx across the brush border of the rat jejunum. Effects of a diet rich in fructose].

PubMed

Crouzoulon, G

1978-10-01

The unidirectional influx (i.e. initial rate of uptake) of D-fructose across the brush border of rat jejunum is a saturable function of concentration, with a Kt of 125 mM, which implicates a carrier mechanism. This mechanism appears to be very specific for fructose in view of the lack of influx inhibition observed in the presence of large concentrations of the sugars or polyols, D-glucose, D-galactose, D-mannose, D-xylose, L-sorbose, D-tagatose, sorbitol or mannitol. D-Fructose uptake is inhibited by incubation, preceded by a 30-min preincubation in the same inhibitory conditions, in the absence of Na, or in the presence of metabolic poisons, NaF, 2,4-dinitrophenol, monoiodoacetate. Phloridzin (10-3 M), with or without preincubation, has no effect on uptake. D-Fructose influx is stimulated by fructose feeding, mainly because the augmentation of the number of active sites of transfer: Jmax is increased two-fold, Kt is more weakly affected.

Isolation and properties of an endo-β-mannanase-producing Bacillus sp. LX114 capable of degrading guar gum.

PubMed

Jiang, Baohang; Sun, Zhen; Hou, Yingmin; Yang, Lan; Yang, Fan; Chen, Xiaoyi; Li, Xianzhen

2016-07-03

Endo-β-mannanase, catalyzing the random hydrolysis of β-1,4-mannosidic linkage in the backbone of (hetero) mannan, can increase feed conversion efficiency of animal feed or form functional mannanooligosaccharides. In this study, a gram-positive, straight-rod, facultative anaeorobic bacterium producing endo-β-mannanase was isolated from soil sample. The isolate only fermented glucose, galactose, sorbose, and raffinose to acid. The test in hydrogen sulfide production was positive. Combining the data acquired from phenotypic analysis and phylogenetic analysis based on 16S rRNA gene sequences, this strain presumably represented a novel species of the genus Bacillus and was designated as LX114. The strain LX114 could break down guar gum molecules, leading to a rapid decrease of the viscosity of guar gum solutions. Endo-β-mannanase activity was also detected in the culture supernatant. The isolate LX114 would be useful for potential application in degrading plant cell walls for increasing feed conversion efficiency and formation of functional oligosaccharides.

Synthesis of glycogen from fructose in the presence of elevated levels of glycogen phosphorylase a in rat hepatocytes.

PubMed

Ciudad, C J; Massagué, J; Salavert, A; Guinovart, J J

1980-03-20

Incubation of hepatocytes with glucose promoted the increase in the glycogen synthase (-glucose 6-phosphate/+glucose 6-phosphate) activity ratio, the decrease in the levels of phosphorylase a and a marked increase in the intracellular glycogen level. Incubation with fructose alone promoted the simultaneous activation of glycogen synthase and increase in the levels of phosphorylase a. Strikingly, glycogen deposition occurred in spite of the elevated levels of phosphorylase a. When glucose and fructose were added to the media the activation of glycogen synthase was always higher than when the hexoses were added separately. On the other hand the effects on glycogen phosphorylase were a function of the relative concentrations of both sugars. Inactivation of glycogen phosphorylase occurred when the fructose to glucose ratio was low while activation took place when the ratio was high. The simultaneous presence of glucose and fructose resulted, in all cases, in an enhancement in the deposition of glycogen. The effects described were not limited to fructose as D-glyceraldehyde, dihydroxyacetone, L-sorbose, D-tagatose and sorbitol, compounds metabolically related to fructose, provoked the same behaviour.

Influence of carbohydrates on secondary metabolism in Fusarium avenaceum.

PubMed

Sørensen, Jens Laurids; Giese, Henriette

2013-09-24

Fusarium avenaceum is a widespread pathogen of important crops in the temperate climate zones that can produce many bioactive secondary metabolites, including moniliformin, fusarin C, antibiotic Y, 2-amino-14,16-dimethyloctadecan-3-ol (2-AOD-3-ol), chlamydosporol, aurofusarin and enniatins. Here, we examine the production of these secondary metabolites in response to cultivation on different carbon sources in order to gain insight into the regulation and production of secondary metabolites in F. avenaceum. Seven monosaccharides (arabinose, xylose, fructose, sorbose, galactose, mannose, glucose), five disaccharides (cellobiose, lactose, maltose, sucrose and trehalose) and three polysaccharides (dextrin, inulin and xylan) were used as substrates. Three F. avenaceum strains were used in the experiments. These were all able to grow and produce aurofusarin on the tested carbon sources. Moniliformin and enniatins were produced on all carbon types, except on lactose, which suggest a common conserved regulation mechanism. Differences in the strains was observed for production of fusarin C, 2-AOD-3-ol, chlamydosporol and antibiotic Y, which suggests that carbon source plays a role in the regulation of their biosynthesis.

TM0416, a Hyperthermophilic Promiscuous Nonphosphorylated Sugar Isomerase, Catalyzes Various C5 and C6 Epimerization Reactions

PubMed Central

Shin, Sun-Mi; Cao, Thinh-Phat; Choi, Jin Myung; Kim, Seong-Bo; Lee, Sang-Jae

2017-01-01

ABSTRACT There is currently little information on nonphosphorylated sugar epimerases, which are of potential interest for producing rare sugars. We found a gene (the TM0416 gene) encoding a putative d-tagatose-3-epimerase-related protein from the hyperthermophilic bacterium Thermotoga maritima. We overexpressed the TM0416 gene in Escherichia coli and purified the resulting recombinant protein for detailed characterization. Amino acid sequence alignment and a structural similarity search revealed that TM0416 is a putative nonphosphorylated sugar epimerase. The recombinant enzyme exhibited maximal C-3 epimerization of l-ribulose to l-xylulose at ∼80°C and pH 7 in the presence of 1 mM Mn2+. In addition, this enzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose, with a conversion yield of 20% after 6 h at 80°C. Remarkably, the enzyme catalyzed the isomerization of d-erythrose or d-threose to d-erythrulose significantly, with conversion yields of 71% and 54.5%, respectively, after 6 h at 80°C at pH 7. To further investigate the substrate specificity of TM0416, we determined its crystal structures in complex with divalent metal ions and l-erythrulose at resolutions of 1.5 and 1.6 Å. Detailed inspection of the structural features and biochemical data clearly demonstrated that this metalloenzyme, with a freely accessible substrate-binding site and neighboring hydrophobic residues, exhibits different and promiscuous substrate preferences, compared with its mesophilic counterparts. Therefore, this study suggests that TM0416 can be functionally classified as a novel type of l-ribulose 3-epimerase (R3E) with d-erythrose isomerase activity. IMPORTANCE Rare sugars, which occur naturally in small amounts, have attracted considerable attention in the food and drug industries. However, there is little information on nonphosphorylated sugar epimerases, which might potentially be applied for the production of rare sugars. This study describes the characterization and functional annotation of a putative nonphosphorylated sugar 3-epimerase from a hyperthermophilic bacterium. Furthermore, we determined its crystal structures in complex with divalent metal ions and l-erythrulose, highlighting its metal-dependent, bifunctional, sugar-isomerizing activity. This hyperthermophilic R3E exhibited d-erythrose/d-threose isomerase activity, with structural features near the substrate-binding site distinct from those of its mesophilic counterparts. Moreover, this metalloenzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose at 70°C. Therefore, TM0416 can be functionally classified as a novel type of promiscuous R3E with a potential for the production of rare sugars for the food and pharmaceutical industries. PMID:28258150

TM0416, a Hyperthermophilic Promiscuous Nonphosphorylated Sugar Isomerase, Catalyzes Various C5 and C6 Epimerization Reactions.

PubMed

Shin, Sun-Mi; Cao, Thinh-Phat; Choi, Jin Myung; Kim, Seong-Bo; Lee, Sang-Jae; Lee, Sung Haeng; Lee, Dong-Woo

2017-05-15

There is currently little information on nonphosphorylated sugar epimerases, which are of potential interest for producing rare sugars. We found a gene (the TM0416 gene) encoding a putative d-tagatose-3-epimerase-related protein from the hyperthermophilic bacterium Thermotoga maritima We overexpressed the TM0416 gene in Escherichia coli and purified the resulting recombinant protein for detailed characterization. Amino acid sequence alignment and a structural similarity search revealed that TM0416 is a putative nonphosphorylated sugar epimerase. The recombinant enzyme exhibited maximal C-3 epimerization of l-ribulose to l-xylulose at ∼80°C and pH 7 in the presence of 1 mM Mn 2+ In addition, this enzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose, with a conversion yield of 20% after 6 h at 80°C. Remarkably, the enzyme catalyzed the isomerization of d-erythrose or d-threose to d-erythrulose significantly, with conversion yields of 71% and 54.5%, respectively, after 6 h at 80°C at pH 7. To further investigate the substrate specificity of TM0416, we determined its crystal structures in complex with divalent metal ions and l-erythrulose at resolutions of 1.5 and 1.6 Å. Detailed inspection of the structural features and biochemical data clearly demonstrated that this metalloenzyme, with a freely accessible substrate-binding site and neighboring hydrophobic residues, exhibits different and promiscuous substrate preferences, compared with its mesophilic counterparts. Therefore, this study suggests that TM0416 can be functionally classified as a novel type of l-ribulose 3-epimerase (R3E) with d-erythrose isomerase activity. IMPORTANCE Rare sugars, which occur naturally in small amounts, have attracted considerable attention in the food and drug industries. However, there is little information on nonphosphorylated sugar epimerases, which might potentially be applied for the production of rare sugars. This study describes the characterization and functional annotation of a putative nonphosphorylated sugar 3-epimerase from a hyperthermophilic bacterium. Furthermore, we determined its crystal structures in complex with divalent metal ions and l-erythrulose, highlighting its metal-dependent, bifunctional, sugar-isomerizing activity. This hyperthermophilic R3E exhibited d-erythrose/d-threose isomerase activity, with structural features near the substrate-binding site distinct from those of its mesophilic counterparts. Moreover, this metalloenzyme showed unusually high activity for the epimerization of d-tagatose to d-sorbose at 70°C. Therefore, TM0416 can be functionally classified as a novel type of promiscuous R3E with a potential for the production of rare sugars for the food and pharmaceutical industries. Copyright © 2017 American Society for Microbiology.

Presumptive speciation of Streptococcus bovis and other group D streptococci from human sources by using arginine and pyruvate tests.

PubMed Central

Gross, K C; Houghton, M P; Senterfit, L B

1975-01-01

A simplified method for speciation of group D streptococci is described. A total of 4,156 streptococcal isolates from human clinical material was tested for ability to hydrolyze esculin in the presence of 40% bile, ferment pyruvate, hydrolyze arginine, and grow in media containing 40% bile or 6.5% NaCl. Streptococci which hydrolyzed esculin in 40% bile, but which did not hydrolyze arginine, were also tested for their ability to ferment raffinose or sorbose. Sixty percent (2,503) of the isolates hydrolyzed esculin in the presence of 40% bile and were thus presumptively identified as group D. By application of the other criteria, 84% of these were speciated as Streptococcus faecalis, 7% were speciated as S. faecium, 6% were speciated as S. bovis, 2% were speciated as S. avium, and 1% were not identified. This scheme was shown to be both reliable and practical for use in the diagnostic laboratory. S. avium and S. bovis isolates were characterized, and 18 S. bovis isolates from patients with bacterial endocarditis were compared physiologically with 151 isolates of this species from other sources. PMID:1176592

Expedient synthesis of C-aryl carbohydrates by consecutive biocatalytic benzoin and aldol reactions.

PubMed

Hernández, Karel; Parella, Teodor; Joglar, Jesús; Bujons, Jordi; Pohl, Martina; Clapés, Pere

2015-02-16

The introduction of aromatic residues connected by a C-C bond into the non-reducing end of carbohydrates is highly significant for the development of innovative structures with improved binding affinity and selectivity (e.g., C-aril-sLex). In this work, an expedient asymmetric "de novo" synthetic route to new aryl carbohydrate derivatives based on two sequential stereoselectively biocatalytic carboligation reactions is presented. First, the benzoin reaction of aromatic aldehydes to dimethoxyacetaldehyde is conducted, catalyzed by benzaldehyde lyase from Pseudomonas fluorescens biovar I. Then, the α-hydroxyketones formed are reduced by using NaBH4 yielding the anti diol. After acetal hydrolysis, the aldol addition of dihydroxyacetone, hydroxyacetone, or glycolaldehyde catalyzed by the stereocomplementary D-fructose-6-phosphate aldolase and L-rhamnulose-1-phosphate aldolase is performed. Both aldolases accept unphosphorylated donor substrates, avoiding the need of handling the phosphate group that the dihydroxyacetone phosphate-dependent aldolases require. In this way, 6-C-aryl-L-sorbose, 6-C-aryl-L-fructose, 6-C-aryl-L-tagatose, and 5-C-aryl-L-xylose derivatives are prepared by using this methodology. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Evaluation of carbohydrates in natural and cultured Cordyceps by pressurized liquid extraction and gas chromatography coupled with mass spectrometry.

PubMed

Guan, Jia; Yang, Feng-Qing; Li, Shao-Ping

2010-06-11

Free and polymeric carbohydrates in Cordyceps, a valued edible mushroom and well-known traditional Chinese medicine, were determined using stepwise pressurized liquid extraction (PLE) extraction and GC-MS. Based on the optimized PLE conditions, acid hydrolysis and derivatization, ten monosaccharides, namely rhamnose, ribose, arabinose, xylose, mannose, glucose, galactose, mannitol, fructose and sorbose in 13 samples of natural and cultured Cordyceps were qualitatively and quantitatively analyzed and compared with myo-inositol hexaacetate as internal standard. The results showed that natural C. sinensis contained more than 7.99% free mannitol and a small amount of glucose, while its polysaccharides were usually composed of mannose, glucose and galactose with a molar ratio of 1.00:16.61-3.82:1.60-1.28. However, mannitol in cultured C. sinensis and cultured C. militaris were less than 5.83%, and free glucose was only detected in a few samples, while their polysaccharides were mainly composed of mannose, glucose and galactose with molar ratios of 1.00:3.01-1.09:3.30-1.05 and 1.00:2.86-1.28:1.07-0.78, respectively. Natural and cultured Cordyceps could be discriminated by hierarchical clustering analysis based on its free carbohydrate contents.

UTILIZATION OF CARBOHYDRATES AND POLYHYDRIC ALCOHOLS BY MYCOBACTERIUM TUBERCULOSIS

PubMed Central

Sweeney, Edward E.; Jann, Gregory J.

1962-01-01

Sweeney, Edward E. (University of California, Los Angeles) and Gregory J. Jann. Utilization of carbohydrates and polyhydric alcohols by Mycobacterium tuberculosis. J. Bacteriol. 84:459–465. 1962.—A new procedure, using a massive inoculum and nongrowth basal medium, was employed for testing carbohydrate and polyhydric alcohol utilization by human tubercle bacilli. A positive reaction was represented by acidification of the test medium rather than by growth, which was the criterion for carbohydrate utilization in studies by earlier workers. The new procedure was both more sensitive and more rapid than growth techniques; results were obtained within days, compared to weeks or months required for growth testing. The massive-inoculum technique may be applied to compounds other than carbohydrates and polyhydric alcohols, and is a sensitive means of detecting changes wrought by various chemical and physical agents upon the metabolism of tubercle bacilli. Three H37Rv strains and six strains of human tubercle bacilli freshly isolated from patients were tested with 21 carbohydrates and polyhydric alcohols. All nine strains gave strong positive reactions for glucose and glycerol, and usually weak positive reactions for ribose and sorbose. Five of the nine strains were trehalose positive, and six (all fresh patient isolates) of the nine were mannose positive. PMID:13979661

The role of artificial and natural sweeteners in reducing the consumption of table sugar: A narrative review.

PubMed

Mooradian, Arshag D; Smith, Meridith; Tokuda, Masaaki

2017-04-01

The rapid increase in the prevalence of obesity worldwide has been partially attributed to the overconsumption of added sugars. Recent guidelines call for limiting the consumption of simple sugars to less than 10% of daily caloric consumption. High intensity sweeteners are regulated as food additives and include aspartame, acesulfame-k, neotame, saccharin, sucralose, cyclamate and alitame. Steviol glycosides and Luo Han Guo fruit extracts are high intensity sweeteners that are designated as generally recognized as safe (GRAS). Commonly used non-caloric artificial sweeteners may have unfavorable effect on health including glucose intolerance and failure to cause weight reduction. The nutritive sweeteners include sugar alcohols such as sorbitol, xylitol, lactitol, mannitol, erythritol, trehalose and maltitol. Naturally occurring rare sugars have recently emerged as an alternative category of sweeteners. These monosaccharides and their derivatives are found in nature in small quantities and lack significant calories. This category includes d-allulose (d-psicose), d-tagatose, d-sorbose and d-allose. Limiting consumption of any sweetener may well be the best health advice. Identifying natural sweeteners that have favorable effects on body weight and metabolism may help achieving the current recommendations of restricting simple sugar consumption. Copyright © 2017 European Society for Clinical Nutrition and Metabolism. Published by Elsevier Ltd. All rights reserved.

Fructose and tagatose protect against oxidative cell injury by iron chelation.

PubMed

Valeri, F; Boess, F; Wolf, A; Göldlin, C; Boelsterli, U A

1997-01-01

To further investigate the mechanism by which fructose affords protection against oxidative cell injury, cultured rat hepatocytes were exposed to cocaine (300 microM) or nitrofurantoin (400 microM). Both drugs elicited massively increased lactate dehydrogenase release. The addition of the ketohexoses D-fructose (metabolized via glycolysis) or D-tagatose (poor glycolytic substrate) significantly attenuated cocaine- and nitrofurantoin-induced cell injury, although both fructose and tagatose caused a rapid depletion of ATP and compromised the cellular energy charge. Furthermore, fructose, tagatose, and sorbose all inhibited in a concentration-dependent manner (0-16 mM) luminolenhanced chemiluminescence (CL) in cell homogenates, indicating that these compounds inhibit the iron-dependent reactive oxygen species (ROS)-mediated peroxidation of luminol. Indeed, both Fe2+ and Fe3+ further increased cocaine-stimulated CL, which was markedly quenched following addition of the ketohexoses. The iron-independent formation of superoxide anion radicals (acetylated cytochrome c reduction) induced by the prooxidant drugs remained unaffected by fructose or tagatose. The iron-chelator deferoxamine similarly protected against prooxidant-induced cell injury. In contrast, the nonchelating aldohexoses D-glucose and D-galactose did not inhibit luminol CL nor did they protect against oxidative cell injury. These data indicate that ketohexoses can effectively protect against prooxidant-induced cell injury, independent of their glycolytic metabolism, by suppressing the iron-catalyzed formation of ROS.

Mechanism of activation of glycogen phosphorylase by fructose in the liver. Stimulation of phosphorylase kinase related to the consumption of adenosine triphosphate.

PubMed

Van de Werve, G; Hers, H G

1979-01-15

1. A dose-dependent activation of phosphorylase and consumption of ATP was observed in isolated hepatocytes incubated in the presence of fructose; histone kinase and phosphorylase kinase activities were unchanged at doses of this sugar that were fully effective on phosphorylase. The activation of phosphorylase by fructose was also observed in cells incubated in a Ca2+-free medium as well as in the livers of rats in vivo. 2. In a liver high-speed supernatant, fructose, tagatose and sorbose stimulated the activity of phosphorylase kinase; this effect was dependent on the presence of K+ ions, which are required for the activity of fructokinase; it was accompanied by the transformation of ATP into ADP. In the presence of hexokinase, glucose also stimulated phosphorylase kinase, both in an Na+ or a K+ medium. 3. The activities of partially purified muscle or liver phosphorylase kinase were unchanged in the presence of fructose. 4. Some properties of liver phosphorylase kinase are described, including a high molecular weight and an inhibition at ATP/Mg ratios above 0.5, as well as an effect of ATP concentration on the hysteretic behaviour of this enzyme. 5. The effect of fructose on the activation of phosphorylase is discussed in relation to the comsumption of ATP.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kawakishi, S.; Okumura, J.; Namiki, M.

Effects of gamma -irradiation on the amimo-carbonyl reaction were investigated with the medel system of various sugar-glycine solutions. The mixed solutions of sugar and glycine were irradiatnd at 20 deg C with gamma rays from a /sup 60/Co source under the presence and absence of oxygen. The browning, the increase in absorbance at 420 nm of this solution with heating, was enhanced by irradiation, especially at the initial stage of browning reaction, but the extent of browning depended on the kinds of sugars, and fructose, sorbose, and sucrose were more remarkable than other sugars. Their browning was more enhanced inmore » the basic solution than the case of neutml and acidic solution, and they were also increased with irradiation doses. The browning between irradiated sugar and unirradiated glycine solution was similar to that of irradiated sugar-glycine solution, and therefore, it was assumed that this browning reaction was due to some fraction in the irradiated sugar. On the browning in the system of the irradiated fructose-other amino acid, the cases of histidine and tryptophane were more noticeable. Glycolaldehyde, glyceraldehyde and glucosone, which are known to be produced by gamma -radiolysis of sugars, showed the browning on reaction with glycine, and the last one was also detected in the irradiated fructose solution by paper chromatography. (auth)« less

Crystal structures of D-psicose 3-epimerase from Clostridium cellulolyticum H10 and its complex with ketohexose sugars.

PubMed

Chan, Hsiu-Chien; Zhu, Yueming; Hu, Yumei; Ko, Tzu-Ping; Huang, Chun-Hsiang; Ren, Feifei; Chen, Chun-Chi; Ma, Yanhe; Guo, Rey-Ting; Sun, Yuanxia

2012-02-01

D-psicose 3-epimerase (DPEase) is demonstrated to be useful in the bioproduction of D-psicose, a rare hexose sugar, from D-fructose, found plenty in nature. Clostridium cellulolyticum H10 has recently been identified as a DPEase that can epimerize D-fructose to yield D-psicose with a much higher conversion rate when compared with the conventionally used DTEase. In this study, the crystal structure of the C. cellulolyticum DPEase was determined. The enzyme assembles into a tetramer and each subunit shows a (β/α)(8) TIM barrel fold with a Mn(2+) metal ion in the active site. Additional crystal structures of the enzyme in complex with substrates/products (D-psicose, D-fructose, D-tagatose and D-sorbose) were also determined. From the complex structures of C. cellulolyticum DPEase with D-psicose and D-fructose, the enzyme has much more interactions with D-psicose than D-fructose by forming more hydrogen bonds between the substrate and the active site residues. Accordingly, based on these ketohexose-bound complex structures, a C3-O3 proton-exchange mechanism for the conversion between D-psicose and D-fructose is proposed here. These results provide a clear idea for the deprotonation/protonation roles of E150 and E244 in catalysis.

chs-4, a class IV chitin synthase gene from Neurospora crassa.

PubMed

Din, A B; Specht, C A; Robbins, P W; Yarden, O

1996-02-05

In Saccharomyces cerevisiae, most of the cellular chitin is produced by chitin synthase III, which requires the product encoded by the CSD2/CAL1/DIT101/KT12 gene. We have identified, isolated and structurally characterized as CSD2/CAL1/DIT101/KT12 homologue in the filamentous ascomycete Neurospora crassa and have used a "reverse genetics" approach to determine its role in vivo. The yeast gene was used as a heterologous probe for the isolation of a N. crassa gene(designated chs-4) encoding a polypeptide belonging to a class of chitin synthases which we have designated class IV. The predicted polypeptide encoded by this gene is highly similar to those of S. cerevisiae and Candida albicans. N. crassa strains in which chs-4 had been inactivated by the Repeat-Induced point mutation (RIP) process grew and developed in a normal manner under standard growth conditions. However, when grown in the presence of sorbose (a carbon source which induces morphological changes accompanied by elevated chitin content), chitin levels in the chs-4RIP strain were significantly lower than those observed in the wild type. We suggest that CHS4 may serve as an auxiliary enzyme in N. crassa and that, in contrast to yeasts, it is possible that filamentous fungi may have more than one class IV chitin synthase.

Yeast succession in the Amazon fruit Parahancornia amapa as resource partitioning among Drosophila spp.

PubMed Central

Morais, P B; Martins, M B; Klaczko, L B; Mendonça-Hagler, L C; Hagler, A N

1995-01-01

The succession of yeasts colonizing the fallen ripe amapa fruit, from Parahancornia amapa, was examined. The occupation of the substrate depended on both the competitive interactions of yeast species, such as the production of killer toxins, and the selective dispersion by the drosophilid guild of the amapa fruit. The yeast community associated with this Amazon fruit differed from those isolated from other fruits in the same forest. The physiological profile of these yeasts was mostly restricted to the assimilation of a few simple carbon sources, mainly L-sorbose, D-glycerol, DL-lactate, cellobiose, and salicin. Common fruit-associated yeasts of the genera Kloeckera and Hanseniaspora, Candida guilliermondii, and Candida krusei colonized fruits during the first three days after the fruit fell. These yeasts were dispersed and served as food for the invader Drosophila malerkotliana. The resident flies of the Drosophila willistoni group fed selectively on patches of yeasts colonizing fruits 3 to 10 days after the fruit fell. The killer toxin-producing yeasts Pichia kluyveri var. kluyveri and Candida fructus were probably involved in the exclusion of some species during the intermediate stages of fruit deterioration. An increase in pH, inhibiting toxin activity and the depletion of simple sugars, may have promoted an increase in yeast diversity in the later stages of decomposition. The yeast succession provided a patchy environment for the drosophilids sharing this ephemeral substrate. PMID:8534092

The Oxygenase CAO-1 of Neurospora crassa Is a Resveratrol Cleavage Enzyme

PubMed Central

Díaz-Sánchez, Violeta; F. Estrada, Alejandro; Limón, M. Carmen; Al-Babili, Salim

2013-01-01

The genome of the ascomycete Neurospora crassa encodes CAO-1 and CAO-2, two members of the carotenoid cleavage oxygenase family that target double bonds in different substrates. Previous studies demonstrated the role of CAO-2 in cleaving the C40 carotene torulene, a key step in the synthesis of the C35 apocarotenoid pigment neurosporaxanthin. In this work, we investigated the activity of CAO-1, assuming that it may provide retinal, the chromophore of the NOP-1 rhodopsin, by cleaving β-carotene. For this purpose, we tested CAO-1 activity with carotenoid substrates that were, however, not converted. In contrast and consistent with its sequence similarity to family members that act on stilbenes, CAO-1 cleaved the interphenyl Cα-Cβ double bond of resveratrol and its derivative piceatannol. CAO-1 did not convert five other similar stilbenes, indicating a requirement for a minimal number of unmodified hydroxyl groups in the stilbene background. Confirming its biological function in converting stilbenes, adding resveratrol led to a pronounced increase in cao-1 mRNA levels, while light, a key regulator of carotenoid metabolism, did not alter them. Targeted Δcao-1 mutants were not impaired by the presence of resveratrol, a phytoalexin active against different fungi, which did not significantly affect the growth and development of wild-type Neurospora. However, under partial sorbose toxicity, the Δcao-1 colonies exhibited faster radial growth than control strains in the presence of resveratrol, suggesting a moderate toxic effect of resveratrol cleavage products. PMID:23893079

Highly efficient production of rare sugars D-psicose and L-tagatose by two engineered D-tagatose epimerases.

PubMed

Bosshart, Andreas; Wagner, Nina; Lei, Lei; Panke, Sven; Bechtold, Matthias

2016-02-01

Rare sugars are monosaccharides that do not occur in nature in large amounts. However, many of them demonstrate high potential as low-calorie sweetener, chiral building blocks or active pharmaceutical ingredients. Their production by enzymatic means from broadly abundant epimers is an attractive alternative to synthesis by traditional organic chemical means, but often suffers from low space-time yields and high enzyme costs due to rapid enzyme degradation. Here we describe the detailed characterization of two variants of d-tagatose epimerase under operational conditions that were engineered for high stability and high catalytic activity towards the epimerization of d-fructose to d-psicose and l-sorbose to l-tagatose, respectively. A variant optimized for the production of d-psicose showed a very high total turnover number (TTN) of up to 10(8) catalytic events over a catalyst's lifetime, determined under operational conditions at high temperatures in an enzyme-membrane reactor (EMR). Maximum space-time yields as high as 10.6 kg L(-1) d(-1) were obtained with a small laboratory-scale EMR, indicating excellent performance. A variant optimized for the production of l-tagatose performed less stable in the same setting, but still showed a very good TTN of 5.8 × 10(5) and space-time yields of up to 478 g L(-1) d(-1) . Together, these results confirm that large-scale enzymatic access to rare sugars is feasible. © 2015 Wiley Periodicals, Inc.

Genetic transformation of Neurospora tetrasperma, demonstration of repeat-induced point mutation (RIP) in self-crosses and a screen for recessive RIP-defective mutants.

PubMed Central

Bhat, Ashwin; Tamuli, Ranjan; Kasbekar, Durgadas P

2004-01-01

The pseudohomothallic fungus Neurospora tetrasperma is naturally resistant to the antibiotic hygromycin. We discovered that mutation of its erg-3 (sterol C-14 reductase) gene confers a hygromycin-sensitive phenotype that can be used to select transformants on hygromycin medium by complementation with the N. crassa erg-3+ and bacterial hph genes. Cotransformation of hph with PCR-amplified DNA of other genes enabled us to construct strains duplicated for the amplified DNA. Using transformation we constructed self-fertile strains that were homoallelic for an ectopic erg-3+ transgene and a mutant erg-3 allele at the endogenous locus. Self-crosses of these strains yielded erg-3 mutant ascospores that produced colonies with the characteristic morphology on Vogel's sorbose agar described previously for erg-3 mutants of N. crassa. The mutants were generated by repeat-induced point mutation (RIP), a genome defense process that causes numerous G:C to A:T mutations in duplicated DNA sequences. Homozygosity for novel recessive RIP-deficient mutations was signaled by self-crosses of erg-3-duplication strains that fail to produce erg-3 mutant progeny. Using this assay we isolated a UV-induced mutant with a putative partial RIP defect. RIP-induced mutants were isolated in rid-1 and sad-1, which are essential genes, respectively, for RIP and another genome defense mechanism called meiotic silencing by unpaired DNA. PMID:15280231

Structure and Antitumor and Immunomodulatory Activities of a Water-Soluble Polysaccharide from Dimocarpus longan Pulp

PubMed Central

Meng, Fa-Yan; Ning, Yuan-Ling; Qi, Jia; He, Zhou; Jie, Jiang; Lin, Juan-Juan; Huang, Yan-Jun; Li, Fu-Sen; Li, Xue-Hua

2014-01-01

A new water-soluble polysaccharide (longan polysaccharide 1 (LP1)) was extracted and successfully purified from Dimocarpus longan pulp via diethylaminoethyl (DEAE)-cellulose anion-exchange and Sephacryl S-300 HR gel chromatography. The chemical structure was determined using Infrared (IR), gas chromatography (GC) and nuclear magnetic resonance (NMR) analysis. The results indicated that the molecular weight of the sample was 1.1 × 105 Da. Monosaccharide composition analysis revealed that LP1 was composed of Glc, GalA, Ara and Gal in a molar ratio of 5.39:1.04:0.74:0.21. Structural analysis indicated that LP1 consisted of a backbone of →4)-α-d-Glcp-(1→4)-α-d-GalpA-(1→4)-α-d-Glcp-(1→4)-β-d-Glcp-(1→ units with poly saccharide side chains composed of →2)-β-d-Fruf-(1→2)-l-sorbose-(1→ attached to the O-6 position of the α-d-Glcp residues. In vitro experiments indicated that LP1 had significantly high antitumor activity against SKOV3 and HO8910 tumor cells, with inhibition percentages of 40% and 50%, respectively. In addition, LP1 significantly stimulated the production of the cytokine interferon-γ (IFN-γ), increased the activity of murine macrophages and enhanced B- and T-lymphocyte proliferation. The results of this study demonstrate that LP1 has potential applications as a natural antitumor agent with immunomodulatory activity. PMID:24663085

Purification and Characterization of Pyranose Oxidase from the White Rot Fungus Trametes multicolor

PubMed Central

Leitner, Christian; Volc, Jindrich; Haltrich, Dietmar

2001-01-01

We purified an intracellular pyranose oxidase from mycelial extracts of the white rot fungus Trametes multicolor by using ammonium sulfate fractionation, hydrophobic interaction, ion-exchange chromatography, and gel filtration. The native enzyme has a molecular mass of 270 kDa as determined by equilibrium ultracentrifugation and is composed of four identical 68-kDa subunits as determined by matrix-assisted laser desorption ionization mass spectrometry. Each subunit contains one covalently bound flavin adenine dinucleotide as its prosthetic group. The enzyme oxidizes several aldopyranoses specifically at position C-2, and its preferred electron donor substrates are d-glucose, d-xylose, and l-sorbose. During this oxidation reaction electrons are transferred to oxygen, yielding hydrogen peroxide. In addition, the enzyme catalyzes the two-electron reduction of 1,4-benzoquinone, several substituted benzoquinones, and 2,6-dichloroindophenol, as well as the one-electron reduction of the ABTS [2,2′-azinobis(3-ethylbenzthiazolinesulfonic acid)] cation radical. As judged by the catalytic efficiencies (kcat/Km), some of these quinone electron acceptors are much better substrates for pyranose oxidase than oxygen. The optimum pH of the pyranose oxidase-catalyzed reaction depends strongly on the electron acceptor employed and varies from 4 to 8. It has been proposed that the main metabolic function of pyranose oxidase is as a constituent of the ligninolytic system of white rot fungi that provides peroxidases with H2O2. An additional function could be reduction of quinones, key intermediates that are formed during mineralization of lignin. PMID:11472941

Reduction of selenite to elemental selenium by Enterobacter cloacae SLD1a-1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dungan, R.S.; Frankenberger, W.T. Jr.

1998-11-01

The facultative anaerobic bacterium Enterobacter cloacae strain SLD1a-1 was studied in washed cell suspensions to assess optimal conditions required for the reduction of selenite (SeO{sub 3}{sup 2{minus}}) to elemental selenium (Se{sup 0}). Enterobacter cloacae using glucose (1.4 mM) as an electron donor removed 79% of the added SeO{sub 3}{sup 2{minus}} from solution in 2.5 h. Optimal SeO{sub 3}{sup 2{minus}} reduction occurred at a pH of 6.5 and a temperature of 40 C. Carbohydrate sources arabinose, xylose, and sorbose were found to significantly enhance SeO{sub 3}{sup 2{minus}} reduction over that of glucose. The reduction of SeO{sub 3}{sup 2{minus}} at 7.9 {micro}Mmore » was inhibited by nitrate of levels 1 to 100 times greater, nitrite at levels 5 and 10 times greater, while sulfite at levels of two to four times greater was found to stimulate the reduction of SeO{sub 3}{sup 2{minus}}. Enterobacter cloacae grows on anaerobically incubated plates containing NO{sub 3}{sup {minus}} as the sole terminal electron acceptor and acetate as the electron donor. Use of SeO{sub 3}{sup 2{minus}} as the terminal electron acceptor during anaerobic respiration did not support growth and could only be reduced to Se{sup 0} when NO{sub 3}{sup {minus}} was present.« less

Effect of D-psicose used as sucrose replacer on the characteristics of meringue.

PubMed

O'Charoen, Siwaporn; Hayakawa, Shigeru; Matsumoto, Yoshiki; Ogawa, Masahiro

2014-12-01

Excessive intake of sugar-rich foods leads to metabolic syndrome. D-Psicose (Psi) not commonly found in nature, is noncalorie sweetener with a suppressive effect on the blood glucose level. Thus, Psi has the potential to be utilized as a sucrose (Suc) replacer in sugar-rich foods, including meringue-based confectionery (MBC). In this study, we investigated the effect of Psi on the physical and chemical properties of meringue. Meringue was made by whipping egg white and Suc (at a weight ratio of 1:1) and baking at 93 °C for 2 h. Thirty percent of the total weight of Suc was replaced with D-ketohexoses such as Psi, D-fructose, D-tagatose, and D-sorbose. The meringues containing D-ketohexoses had higher specific volume than the meringue not containing D-ketohexoses (Ct-meringue). Baking of meringue caused differences between Psi and the other D-ketohexose meringues. Meringue containing Psi (P30-meringue) had the highest breaking stress (7.00 × 10(5) N/m(2)) and breaking strain (4.40%), resulting in the crunchiest texture. In addition, P30-meringue also had the highest antioxidant activity (491.84 μM TE/mg-meringue determined by ABTS method) and was the brownest due to a Maillard reaction occurring during baking. The replacement of Suc with Psi improved the characteristics of baked meringue. Thus, Psi was found to be useful in modifying the physical and chemical properties of MBC. © 2014 Institute of Food Technologists®

Characterization of Candida isolates from pediatric burn patients.

PubMed Central

Neely, A N; Odds, F C; Basatia, B K; Holder, I A

1988-01-01

To provide more detailed information about Candida epidemiology and pathogenesis in pediatric burn patients, Candida isolates from 113 patients collected over 3 years were identified at the species level and the serotypes and biotypes of the C. albicans isolates were determined. A total of 85% of the patients were colonized or infected by C. albicans, 18% by C. tropicalis, and 11% by C. parapsilosis. Although colonization or infection often was found at multiple sites and times, 87% of the patients were colonized or infected by only one Candida species or strain; the other 13% showed multiple colonizations or infections, some of which occurred simultaneously at the same site. C. albicans biotyping determined the tolerance of the isolates to pH (pH 1.4) and salt; flucytosine, borate, and safranine resistance; and ability to produce proteinase and assimilate urea, sorbose, and citrate; results are expressed as three-digit numbers. For isolates from three different anatomical sites, the distribution of the nine biotype characteristics was similar in all cases but one. Significantly more fecal than wound or throat isolates were resistant to safranine. Sixty-four different serotype-biotype combinations were found in the 96 patients with C. albicans infections or colonizations. Twenty-nine percent of all C. albicans isolates had the partial biotype -57, while 20 of the 96 patients had specifically serotype B, biotype 557 colonizations or infections. Eleven patients had the B557 infection when admitted; nine patients acquired the yeast in-house. Thirty percent of the C. albicans isolated from 23 adult patients at a nearby hospital also showed the -57 biotype pattern, suggesting that C. albicans isolates expressing this biotype are either extremely prevalent in nature or are more virulent than other C. albicans isolates. PMID:3053771

Structure of a class I tagatose-1,6-bisphosphate aldolase: investigation into an apparent loss of stereospecificity.

PubMed

LowKam, Clotilde; Liotard, Brigitte; Sygusch, Jurgen

2010-07-02

Tagatose-1,6-bisphosphate aldolase from Streptococcus pyogenes is a class I aldolase that exhibits a remarkable lack of chiral discrimination with respect to the configuration of hydroxyl groups at both C3 and C4 positions. The enzyme catalyzes the reversible cleavage of four diastereoisomers (fructose 1,6-bisphosphate (FBP), psicose 1,6-bisphosphate, sorbose 1,6-bisphosphate, and tagatose 1,6-bisphosphate) to dihydroxyacetone phosphate (DHAP) and d-glyceraldehyde 3-phosphate with high catalytic efficiency. To investigate its enzymatic mechanism, high resolution crystal structures were determined of both native enzyme and native enzyme in complex with dihydroxyacetone-P. The electron density map revealed a (alpha/beta)(8) fold in each dimeric subunit. Flash-cooled crystals of native enzyme soaked with dihydroxyacetone phosphate trapped a covalent intermediate with carbanionic character at Lys(205), different from the enamine mesomer bound in stereospecific class I FBP aldolase. Structural analysis indicates extensive active site conservation with respect to class I FBP aldolases, including conserved conformational responses to DHAP binding and conserved stereospecific proton transfer at the DHAP C3 carbon mediated by a proximal water molecule. Exchange reactions with tritiated water and tritium-labeled DHAP at C3 hydrogen were carried out in both solution and crystalline state to assess stereochemical control at C3. The kinetic studies show labeling at both pro-R and pro-S C3 positions of DHAP yet detritiation only at the C3 pro-S-labeled position. Detritiation of the C3 pro-R label was not detected and is consistent with preferential cis-trans isomerism about the C2-C3 bond in the carbanion as the mechanism responsible for C3 epimerization in tagatose-1,6-bisphosphate aldolase.

X-ray structures of the Pseudomonas cichorii D-tagatose 3-epimerase mutant form C66S recognizing deoxy sugars as substrates.

PubMed

Yoshida, Hiromi; Yoshihara, Akihide; Ishii, Tomohiko; Izumori, Ken; Kamitori, Shigehiro

2016-12-01

Pseudomonas cichorii D-tagatose 3-epimerase (PcDTE), which has a broad substrate specificity, efficiently catalyzes the epimerization of not only D-tagatose to D-sorbose but also D-fructose to D-psicose (D-allulose) and also recognizes the deoxy sugars as substrates. In an attempt to elucidate the substrate recognition and catalytic reaction mechanisms of PcDTE for deoxy sugars, the X-ray structures of the PcDTE mutant form with the replacement of Cys66 by Ser (PcDTE_C66S) in complexes with deoxy sugars were determined. These X-ray structures showed that substrate recognition by the enzyme at the 1-, 2-, and 3-positions is responsible for enzymatic activity and that substrate-enzyme interactions at the 4-, 5-, and 6-positions are not essential for the catalytic reaction of the enzyme leading to the broad substrate specificity of PcDTE. They also showed that the epimerization site of 1-deoxy 3-keto D-galactitol is shifted from C3 to C4 and that 1-deoxy sugars may bind to the catalytic site in the inhibitor-binding mode. The hydrophobic groove that acts as an accessible surface for substrate binding is formed through the dimerization of PcDTE. In PcDTE_C66S/deoxy sugar complex structures, bound ligand molecules in both the linear and ring forms were detected in the hydrophobic groove, while bound ligand molecules in the catalytic site were in the linear form. This result suggests that the sugar-ring opening of a substrate may occur in the hydrophobic groove and also that the narrow channel of the passageway to the catalytic site allows a substrate in the linear form to pass through.

Discovery of a Kojibiose Phosphorylase in Escherichia coli K-12.

PubMed

Mukherjee, Keya; Narindoshvili, Tamari; Raushel, Frank M

2018-05-15

The substrate profiles for three uncharacterized enzymes (YcjM, YcjT, and YcjU) that are expressed from a cluster of 12 genes ( ycjM-W and ompG) of unknown function in Escherichia coli K-12 were determined. Through a comprehensive bioinformatic and steady-state kinetic analysis, the catalytic function of YcjT was determined to be kojibiose phosphorylase. In the presence of saturating phosphate and kojibiose (α-(1,2)-d-glucose-d-glucose), this enzyme catalyzes the formation of d-glucose and β-d-glucose-1-phosphate ( k cat = 1.1 s -1 , K m = 1.05 mM, and k cat / K m = 1.12 × 10 3 M -1 s -1 ). Additionally, it was also shown that in the presence of β-d-glucose-1-phosphate, YcjT can catalyze the formation of other disaccharides using 1,5-anhydro-d-glucitol, l-sorbose, d-sorbitol, or l-iditol as a substitute for d-glucose. Kojibiose is a component of cell wall lipoteichoic acids in Gram-positive bacteria and is of interest as a potential low-calorie sweetener and prebiotic. YcjU was determined to be a β-phosphoglucomutase that catalyzes the isomerization of β-d-glucose-1-phosphate ( k cat = 21 s -1 , K m = 18 μM, and k cat / K m = 1.1 × 10 6 M -1 s -1 ) to d-glucose-6-phosphate. YcjU was also shown to exhibit catalytic activity with β-d-allose-1-phosphate, β-d-mannose-1-phosphate, and β-d-galactose-1-phosphate. YcjM catalyzes the phosphorolysis of α-(1,2)-d-glucose-d-glycerate with a k cat = 2.1 s -1 , K m = 69 μM, and k cat / K m = 3.1 × 10 4 M -1 s -1 .

Structure of a Class I Tagatose-1,6-bisphosphate Aldolase

PubMed Central

LowKam, Clotilde; Liotard, Brigitte; Sygusch, Jurgen

2010-01-01

Tagatose-1,6-bisphosphate aldolase from Streptococcus pyogenes is a class I aldolase that exhibits a remarkable lack of chiral discrimination with respect to the configuration of hydroxyl groups at both C3 and C4 positions. The enzyme catalyzes the reversible cleavage of four diastereoisomers (fructose 1,6-bisphosphate (FBP), psicose 1,6-bisphosphate, sorbose 1,6-bisphosphate, and tagatose 1,6-bisphosphate) to dihydroxyacetone phosphate (DHAP) and d-glyceraldehyde 3-phosphate with high catalytic efficiency. To investigate its enzymatic mechanism, high resolution crystal structures were determined of both native enzyme and native enzyme in complex with dihydroxyacetone-P. The electron density map revealed a (α/β)8 fold in each dimeric subunit. Flash-cooled crystals of native enzyme soaked with dihydroxyacetone phosphate trapped a covalent intermediate with carbanionic character at Lys205, different from the enamine mesomer bound in stereospecific class I FBP aldolase. Structural analysis indicates extensive active site conservation with respect to class I FBP aldolases, including conserved conformational responses to DHAP binding and conserved stereospecific proton transfer at the DHAP C3 carbon mediated by a proximal water molecule. Exchange reactions with tritiated water and tritium-labeled DHAP at C3 hydrogen were carried out in both solution and crystalline state to assess stereochemical control at C3. The kinetic studies show labeling at both pro-R and pro-S C3 positions of DHAP yet detritiation only at the C3 pro-S-labeled position. Detritiation of the C3 pro-R label was not detected and is consistent with preferential cis-trans isomerism about the C2–C3 bond in the carbanion as the mechanism responsible for C3 epimerization in tagatose-1,6-bisphosphate aldolase. PMID:20427286

Cytoprotection by fructose and other ketohexoses during bile salt-induced apoptosis of hepatocytes.

PubMed

Zeid, I M; Bronk, S F; Fesmier, P J; Gores, G J

1997-01-01

Toxic bile salts cause hepatocyte necrosis at high concentrations and apoptosis at lower concentrations. Although fructose prevents bile salt-induced necrosis, the effect of fructose on bile salt-induced apoptosis is unclear. Our aim was to determine if fructose also protects against bile salt-induced apoptosis. Fructose inhibited glycochenodeoxycholate (GCDC)-induced apoptosis in a concentration-dependent manner with a maximum inhibition of 72% +/- 10% at 10 mmol/L. First, we determined if fructose inhibited apoptosis by decreasing adenosine triphosphate (ATP) and intracellular pH (pHi). Although fructose decreased ATP to <25% of basal values, oligomycin (an ATP synthase inhibitor) did not inhibit apoptosis despite decreasing ATP to similar values. Fructose (10 mmol/L) decreased intracellular pH (pHi) by 0.2 U. However, extracellular acidification (pH 6.8), which decreased hepatocyte pHi 0.35 U and is known to inhibit necrosis, actually potentiated apoptosis 1.6-fold. Fructose cytoprotection also could not be explained by induction of bcl-2 transcription or metal chelation. Because we could not attribute fructose cytoprotection to metabolic effects, alterations in the expression of bcl-2, or metal chelation, we next determined if the poorly metabolized ketohexoses, tagatose and sorbose, also inhibited apoptosis; unexpectedly, both ketohexoses inhibited apoptosis. Because bile salt-induced apoptosis and necrosis are inhibited by fructose, these data suggest that similar processes initiate bile salt-induced hepatocyte necrosis and apoptosis. In contrast, acidosis, which inhibits necrosis, potentiates apoptosis. Thus, ketohexose-sensitive pathways appear to initiate both bile salt-induced cell apoptosis and necrosis, whereas dissimilar, pH-sensitive, effector mechanisms execute these two different cell death processes.

Metabolomic profiles of arsenic (+3 oxidation state) methyltransferase knockout mice: Effect of sex and arsenic exposure

PubMed Central

Huang, Madelyn C.; Douillet, Christelle; Su, Mingming; Zhou, Kejun; Wu, Tao; Chen, Wenlian; Galanko, Joseph A.; Drobná, Zuzana; Saunders, R. Jesse; Martin, Elizabeth; Fry, Rebecca C.; Jia, Wei; Stýblo, Miroslav

2016-01-01

Arsenic (+3 oxidation state) methyltransferase (As3mt) is the key enzyme in the pathway for methylation of inorganic arsenic (iAs). Altered As3mt expression and AS3MT polymorphism have been linked to changes in iAs metabolism and in susceptibility to iAs toxicity in laboratory models and in humans. As3mt-knockout mice have been used to study the association between iAs metabolism and adverse effects of iAs exposure. However, little is known about systemic changes in metabolism of these mice and how these changes lead to their increased susceptibility to iAs toxicity. Here, we compared plasma and urinary metabolomes of male and female wild-type (WT) and As3mt-KO (KO) C57BL6 mice and examined metabolomic shifts associated with iAs exposure in drinking water. Surprisingly, exposure to 1 ppm As elicited only small changes in the metabolite profiles of either WT or KO mice. In contrast, comparisons of KO mice with WT mice revealed significant differences in plasma and urinary metabolites associated with lipid (phosphatidylcholines, cytidine, acyl-carnitine), amino acid (hippuric acid, acetylglycine, urea), and carbohydrate (L-sorbose, galactonic acid, gluconic acid) metabolism. Notably, most of these differences were sex-specific. Sex-specific differences were also found between WT and KO mice in plasma triglyceride and lipoprotein cholesterol levels. Some of the differentially changed metabolites (phosphatidylcholines, carnosine, and sarcosine) are substrates or products of reactions catalyzed by other methyltransferases. These results suggest that As3mt KO alters major metabolic pathways in a sex-specific manner, independent of iAs treatment, and that As3mt may be involved in other cellular processes beyond iAs methylation. PMID:26883664

Molecular and crystal structure and the Hirshfeld surface analysis of 1-amino-1-deoxy-α-D-sorbopyranose and 1-amino-1-deoxy-α-D-psicopyranose ("D-sorbosamine" and "D-psicosamine") derivatives

NASA Astrophysics Data System (ADS)

Mossine, Valeri V.; Barnes, Charles L.; Mawhinney, Thomas P.

2018-05-01

Sorbosamine and psicosamine are the last two 1-amino-1-deoxy-hexuloses for which no structural data were available. We report on a13C NMR and a single crystal X-ray diffraction study of 1-deoxy-1-(N-methylphenylamino)-D-sorbose (1) and 1-deoxy-1-(N-methylphenylamino)-D-psicose (2). In solutions, both aminosugars are conformationally unstable and establish equilibria, with 90.7% α-pyranose, 3.8% α-furanose, 1.0% β-pyranose, 0.5% β-furanose, and 4.0% acyclic keto form for 1 and 32.4% α-furanose, 27.2% α-pyranose, 21.0% β-pyranose, 9.1% β-furanose, and 11.0% acyclic keto form for 2. X-ray diffraction data provided detailed structural information on 1 and 2 in the α-pyranose form. Both molecules adopt the 5C2 ring conformations, the bond distances and valence angles compare well with respective pyranose structures. All hydroxyl groups in crystal structures of both 1 and 2 participate in two-dimensional hydrogen bonding networks, the H-bonding pattern in 1 is dominated by co-crystallized water molecules. The Hirshfeld surface analysis revealed a significant contribution of non- or weakly polar interactions to the packing forces for both molecules, with crystal structure of 2 featuring short H⋯H contacts. Other structural features found in 2 are a significant planarity of the tertiary amino group (the pyramid heights are 0.127 Å in 2 vs 0.231 Å in 1), a concomitant non-involvement of the amine nitrogen in heteroatom contacts, and a unique anti-periplanar conformation around the C1sbnd C2 bond.

Comparative genome analysis of the candidate functional starter culture strains Lactobacillus fermentum 222 and Lactobacillus plantarum 80 for controlled cocoa bean fermentation processes.

PubMed

Illeghems, Koen; De Vuyst, Luc; Weckx, Stefan

2015-10-12

Lactobacillus fermentum 222 and Lactobacillus plantarum 80, isolates from a spontaneous Ghanaian cocoa bean fermentation process, proved to be interesting functional starter culture strains for cocoa bean fermentations. Lactobacillus fermentum 222 is a thermotolerant strain, able to dominate the fermentation process, thereby converting citrate and producing mannitol. Lactobacillus plantarum 80 is an acid-tolerant and facultative heterofermentative strain that is competitive during cocoa bean fermentation processes. In this study, whole-genome sequencing and comparative genome analysis was used to investigate the mechanisms of these strains to dominate the cocoa bean fermentation process. Through functional annotation and analysis of the high-coverage contigs obtained through 454 pyrosequencing, plantaricin production was predicted for L. plantarum 80. For L. fermentum 222, genes encoding a complete arginine deiminase pathway were attributed. Further, in-depth functional analysis revealed the capacities of these strains associated with carbohydrate and amino acid metabolism, such as the ability to use alternative external electron acceptors, the presence of an extended pyruvate metabolism, and the occurrence of several amino acid conversion pathways. A comparative genome sequence analysis using publicly available genome sequences of strains of the species L. plantarum and L. fermentum revealed unique features of both strains studied. Indeed, L. fermentum 222 possessed genes encoding additional citrate transporters and enzymes involved in amino acid conversions, whereas L. plantarum 80 is the only member of this species that harboured a gene cluster involved in uptake and consumption of fructose and/or sorbose. In-depth genome sequence analysis of the candidate functional starter culture strains L. fermentum 222 and L. plantarum 80 revealed their metabolic capacities, niche adaptations and functionalities that enable them to dominate the cocoa bean fermentation process. Further, these results offered insights into the cocoa bean fermentation ecosystem as a whole and will facilitate the selection of appropriate starter culture strains for controlled cocoa bean fermentation processes.

Carbohydrates in thermophile metabolism: calculation of the standard molal thermodynamic properties of aqueous pentoses and hexoses at elevated temperatures and pressures

NASA Astrophysics Data System (ADS)

Amend, Jan P.; Plyasunov, Andrey V.

2001-11-01

Experimental thermodynamic data for aqueous organic compounds can be combined with the revised Helgeson-Kirkham-Flowers (HKF) equations of state to generate parameters that can be used to estimate standard molal properties as functions of temperature and pressure. In this study, we regressed thermodynamic data for aqueous carbohydrates at temperatures up to 393 K reported in the literature to permit the calculation of the apparent standard molal Gibbs free energies and enthalpies of formation (ΔGo and ΔHo, respectively) and the standard molal entropies (S2o), heat capacities (CP,2o), and volumes (V2o) to 423 K and several hundred MPa of aqueous C5 aldoses (ribose, arabinose, xylose, lyxose) and C5 ketoses (ribulose, xylulose) as well as C6 aldoses (glucose, mannose, galactose) and C6 ketoses (fructose, sorbose). Values of ΔGo for these 11 aqueous carbohydrates are given as a function of temperature at the saturated water vapor pressure (PSAT) and at 50 MPa. Values of ΔGo for aqueous glucose are then combined with those of other aqueous organic and inorganic compounds to calculate values of the standard molal Gibbs free energies of 13 fermentation and respiration reactions (ΔGro) known or likely to be carried out by thermophilic microorganisms. Finally, values of the overall Gibbs free energies of these reactions (ΔGr) are calculated at the temperature, pressure, and chemical composition that obtain in the hydrothermal fluids of Vulcano Island, southern Italy, a site that is widely known for its tremendous diversity of organisms able to live at high temperatures. At likely activities of aqueous glucose, it is shown that thermophiles in the hot springs of Vulcano at 373 K and ∼0.1 MPa can gain between 400 and 3000 kJ per mole of glucose fermented or respired.

Streptococcus caviae sp. nov., isolated from guinea pig faecal samples.

PubMed

Palakawong Na Ayudthaya, Susakul; Hilderink, Loes J; Oost, John van der; Vos, Willem M de; Plugge, Caroline M

2017-05-01

A novel cellobiose-degrading and lactate-producing bacterium, strain Cavy grass 6T, was isolated from faecal samples of guinea pigs (Cavia porcellus). Cells of the strain were ovalshaped, non-motile, non-spore-forming, Gram-stain-positive and facultatively anaerobic. The strain gr at 25-40 °C (optimum 37 °C) and pH 4.5-9.5 (optimum 8.0). Phylogenetic analysis based on 16S rRNA gene sequences showed that strain Cavy grass 6T belongs to the genus Streptococcus with its closest relative being Streptococcus devriesei CCUG 47155T with only 96.5 % similarity. Comparing strain Cavy grass 6T and Streptococcus devriesei CCUG 47155T, average nucleotide identity and level of digital DNA-DNA hybridization dDDH were only 86.9 and 33.3 %, respectively. Housekeeping genes groEL and gyrA were different between strain Cavy grass 6T and other streptococci. The G+C content of strain Cavy grass 6T was 42.6±0.3 mol%. The major (>10 %) cellular fatty acids of strain Cavy grass 6T were C16:0, C20 : 1ω9c and summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c). Strain Cavy grass 6T ferment a range of plant mono- and disaccharides as well as polymeric carbohydrates, including cellobiose, dulcitol, d-glucose, maltose, raffinose, sucrose, l-sorbose, trehalose, inulin and dried grass extract, to lactate, formate, acetate and ethanol. Based on phylogenetic and physiological characteristics, Cavy grass 6T can be distinguished from other members of the genus Streptococcus. Therefore, a novel species of the genus Streptococcus, family Streptococcaceae, order Lactobacillales is proposed, Streptococcuscaviae sp. nov. (type strain Cavy grass 6T=TISTR 2371T=DSM 102819T).

Quick identification of acetic acid bacteria based on nucleotide sequences of the 16S-23S rDNA internal transcribed spacer region and of the PQQ-dependent alcohol dehydrogenase gene.

PubMed

Trcek, Janja

2005-10-01

Acetic acid bacteria (AAB) are well known for oxidizing different ethanol-containing substrates into various types of vinegar. They are also used for production of some biotechnologically important products, such as sorbose and gluconic acids. However, their presence is not always appreciated since certain species also spoil wine, juice, beer and fruits. To be able to follow AAB in all these processes, the species involved must be identified accurately and quickly. Because of inaccuracy and very time-consuming phenotypic analysis of AAB, the application of molecular methods is necessary. Since the pairwise comparison among the 16S rRNA gene sequences of AAB shows very high similarity (up to 99.9%) other DNA-targets should be used. Our previous studies showed that the restriction analysis of 16S-23S rDNA internal transcribed spacer region is a suitable approach for quick affiliation of an acetic acid bacterium to a distinct group of restriction types and also for quick identification of a potentially novel species of acetic acid bacterium (Trcek & Teuber 2002; Trcek 2002). However, with the exception of two conserved genes, encoding tRNAIle and tRNAAla, the sequences of 16S-23S rDNA are highly divergent among AAB species. For this reason we analyzed in this study a gene encoding PQQ-dependent ADH as a possible DNA-target. First we confirmed the expression of subunit I of PQQ-dependent ADH (AdhA) also in Asaia, the only genus of AAB which exhibits little or no ADH-activity. Further we analyzed the partial sequences of adhA among some representative species of the genera Acetobacter, Gluconobacter and Gluconacetobacter. The conserved and variable regions in these sequences made possible the construction of A. acetispecific oligonucleotide the specificity of which was confirmed in PCR-reaction using 45 well-defined strains of AAB as DNA-templates. The primer was also successfully used in direct identification of A. aceti from home made cider vinegar as well as for revealing the misclassification of strain IFO 3283 into the species A. aceti.

Escherichia coli O26 in feedlot cattle: fecal prevalence, isolation, characterization, and effects of an E. coli O157 vaccine and a direct-fed microbial.

PubMed

Paddock, Zac D; Renter, David G; Cull, Charley A; Shi, Xiarong; Bai, Jianfa; Nagaraja, Tiruvoor G

2014-03-01

Escherichia coli O26 is second only to O157 in causing foodborne, Shiga toxin-producing E. coli (STEC) infections. Our objectives were to determine fecal prevalence and characteristics of E. coli O26 in commercial feedlot cattle (17,148) that were enrolled in a study to evaluate an E. coli O157:H7 siderophore receptor and porin (SRP(®)) vaccine (VAC) and a direct-fed microbial (DFM; 10(6) colony-forming units [CFU]/animal/day of Lactobacillus acidophilus and 10(9) CFU/animal/day of Propionibacterium freudenreichii). Cattle were randomly allocated to 40 pens within 10 complete blocks; pens were randomly assigned to control, VAC, DFM, or VAC+DFM treatments. Vaccine was administered on days 0 and 21, and DFM was fed throughout the study. Pen-floor fecal samples (30/pen) were collected weekly for the last 4 study weeks. Samples were enriched in E. coli broth and subjected to a multiplex polymerase chain reaction (PCR) designed to detect O26-specific wzx gene and four major virulence genes (stx1, stx2, eae, and ehxA) and to a culture-based procedure that involved immunomagnetic separation and plating on MacConkey agar. Ten presumptive E. coli colonies were randomly picked, pooled, and tested by the multiplex PCR. Pooled colonies positive for O26 serogroup were streaked on sorbose MacConkey agar, and 10 randomly picked colonies per sample were tested individually by the multiplex PCR. The overall prevalence of E. coli O26 was higher (p<0.001) by the culture-based method compared to the PCR assay (22.7 versus 10.5%). The interventions (VAC and or DFM) had no impact on fecal shedding of O26. Serogroup O26 was recovered in pure culture from 23.9% (260 of 1089) of O26 PCR-positive pooled colonies. Only 7 of the 260 isolates were positive for the stx gene and 90.1% of the isolates possessed an eaeβ gene that codes for intimin subtype β, but not the bfpA gene, which codes for bundle-forming pilus. Therefore, the majority of the O26 recovered from feedlot cattle feces was atypical enteropathogenic E. coli, and not STEC.

Crystal structures of D-tagatose 3-epimerase from Pseudomonas cichorii and its complexes with D-tagatose and D-fructose.

PubMed

Yoshida, Hiromi; Yamada, Mitsugu; Nishitani, Takeyori; Takada, Goro; Izumori, Ken; Kamitori, Shigehiro

2007-11-23

Pseudomonas cichoriiid-tagatose 3-epimerase (P. cichoriid-TE) can efficiently catalyze the epimerization of not only d-tagatose to d-sorbose, but also d-fructose to d-psicose, and is used for the production of d-psicose from d-fructose. The crystal structures of P. cichoriid-TE alone and in complexes with d-tagatose and d-fructose were determined at resolutions of 1.79, 2.28, and 2.06 A, respectively. A subunit of P. cichoriid-TE adopts a (beta/alpha)(8) barrel structure, and a metal ion (Mn(2+)) found in the active site is coordinated by Glu152, Asp185, His211, and Glu246 at the end of the beta-barrel. P. cichoriid-TE forms a stable dimer to give a favorable accessible surface for substrate binding on the front side of the dimer. The simulated omit map indicates that O2 and O3 of d-tagatose and/or d-fructose coordinate Mn(2+), and that C3-O3 is located between carboxyl groups of Glu152 and Glu246, supporting the previously proposed mechanism of deprotonation/protonation at C3 by two Glu residues. Although the electron density is poor at the 4-, 5-, and 6-positions of the substrates, substrate-enzyme interactions can be deduced from the significant electron density at O6. The O6 possibly interacts with Cys66 via hydrogen bonding, whereas O4 and O5 in d-tagatose and O4 in d-fructose do not undergo hydrogen bonding to the enzyme and are in a hydrophobic environment created by Phe7, Trp15, Trp113, and Phe248. Due to the lack of specific interactions between the enzyme and its substrates at the 4- and 5-positions, P. cichoriid-TE loosely recognizes substrates in this region, allowing it to efficiently catalyze the epimerization of d-tagatose and d-fructose (C4 epimer of d-tagatose) as well. Furthermore, a C3-O3 proton-exchange mechanism for P. cichoriid-TE is suggested by X-ray structural analysis, providing a clear explanation for the regulation of the ionization state of Glu152 and Glu246.