Structure Carbohydrate Research Articles

Paenibacillus mellifer sp. nov., isolated from gut of the honey bee Apis mellifera.

An isolate, designated MBLB2552T, was isolated from the gut of the honey bees (Apis mellifera L.) and identified as a member of the genus Paenibacillus based on the sequences of the 16S rRNA gene sequences. The most closely related species to strain MBLB2552T were Paenibacillus timonensis 2301032T, Paenibacillus barengoltzii NBRC 101215T, and Paenibacillus macerans IAM 12467T, with similarity values of 98.1, 97.21 and 97.0%, respectively, based on 16S rRNA gene sequences. The genome size and G + C content of MBLB2552T were 5.2Mb and 52.4%, respectively. The Ortho average nucleotide identity (OrthoANI) and in silico DNA-DNA hybridization (isDDH) values between strain MBLB2552T and the type strains of the closest species were below the species delineation threshold. Comparative genomic analysis showed that most core POGs of strain MBLB2552T and other related taxa were related to translation, ribosomal structure and biogenesis (J) and carbohydrate metabolism in the COG category and KEGG pathways, respectively. Strain MBLB2552T was Gram stain-positive, spore-forming, rod-shaped, facultative anaerobic, motile, and grew at 20‒45°C in 0‒2% (w/v) NaCl at pH 6.0‒9.0. The major polar lipids identified were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylserine, unidentified polar lipids, and an unidentified glycolipid. We propose that strain MBLB2552T represents the type strain of the genus Paenibacillus and its name Paenibacillus mellifer sp. nov. is proposed. The type of strain was MBLB2552T (= JCM 35371T = KCTC 43386T).

Structural analysis of light-colored separated lignin (lignocresol) and its antioxidant properties

The use of lignin is limited by its heterogeneity and complexity. This study was processed using different methods to obtain spruce milled wood lignin (SMWL), spruce kraft lignin (SKL), and spruce lignocresol (SLC) for comparative analysis of the structure and antioxidant activity. SMWL has a complete softwood lignin side-chains structure and lignin carbohydrate complexes. SKL contains fewer ether bonds, while more conjugate structures and condensed structures contribute to the color. However, the α-position of the lignin side chain eliminates most of the hydroxyl and ether bonds (β-O-4/α-OH, phenylcoumaran, and dibenzodioxocine structure) and effectively grafts p-cresol in the phase separation reaction. It not only inhibits the self-condensation of lignin, but also forms the 1,1-diarylpropane unit while protecting β-O-4 linkages from not breaking. Importantly, SLC has few conjugate structures that result in the lightest color among all lignin isolated. Besides, SLC has a high yield and contains trace carbohydrates, indicating that the phase separation method can achieve great amounts of purity separated lignin. The antioxidant activity of lignin was evaluated, results show that 85% of the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals were scavenged at the end of 60 min. Owing to its unique color, structural properties, and continuous antioxidant activity, SLC has the potential to manufacture antioxidant cosmetics.

EXPLORING THE METABOLICALLY ACTIVE RUMEN MICROBIOTA AND ITS FIBROLYTIC POTENTIAL IN CROSSBRED CATTLE FED ON FIBROUS DIET THROUGH METATRANSCRIPTOMICS

The rumen wharves conglomerate of symbiotic fibrolytic microflora with the potential to produce repertoire of carbohydrate-active enzymes to elicit the degradation of recalcitrant plant lignocellulosic complex into simpler forms that can be utilized by the host. A metatranscriptomics approach was adopted to explore the metabolically active microbiota and the transcripts involved in the deconstruction of lignocellulose in Indian crossbred cattle rumen. The experimental animals were fed with mixed diet for a period of twenty-one days and the total RNA was isolated from the rumen digesta using a modified total RNA extraction protocol. After mRNA enrichment, the cDNA libraries were subjected to sequencing under Illumina HiSeq platform and generated about 2.89 GB of raw sequences. The contig assembly was accomplished using two different tools and the redundant sequences were removed before further processing. A total of 133,930 orfs were predicted from the assembled contigs using MetaGeneMark tool and the taxonomic affiliation of orfs were achieved using MEGAN 6.0 Community Edition. A total of 17 bacterial, 5 eukaryotic and 1 archaeal phylum were identified from rumen metatranscriptomic dataset. Phylum Bacteroidetes were acknowledged as the most abundant and metabolically active rumen bacterial populations in cattle rumen with 36,414 orfs corresponding to it, followed by Firmicutes with 6,349 orfs. Further, MEGAN annotation of metatranscriptomic data at species level revealed that Prevotella brevis and Prevotella ruminicola as the most metabolically active bacterial populations representing 4,022 and 2,725 orfs each. The functional annotation of metatranscriptomic data using COG database revealed that a large number of transcripts (~16%) were corresponding to translation, ribosomal structure and biogenesis and carbohydrate transport and metabolism (15%). dbCAN annotation of rumen metatranscriptomic data identified and classified 5,267 transcripts belonging to 168 CAZyme families (GH-52%, GT-26%, CBM-14%, CE-6%, PL-2% and AA-0%). The microbial community analysis of the CAZyme encoding transcripts using M5NR database revealed that a significant proportion of CAZymes were contributed by genera Prevotella (37%) and Bacteroides (21%). The cattle rumen microbiome metatranscriptome analysis presented in this study facilitated the detection of large number of transcripts encoding diverse CAZymes that actively take part in the hydrolysis of plant lignocellulose complex that may be useful for improving livestock and biofuel generation.

Leaf Structural Carbohydrate Decreased for Pinus thunbergii along Coast–Inland Gradients

Although photosynthesis (carbohydrate production) decreases under wind load, it is unclear how carbohydrate categories allocation changes. We determined the leaf morphology (specific leaf area (SLA), needle thickness), anatomy (cuticle thickness, epidermal thickness), photosynthesis (effective quantum yield of Photosystem II (Y(II)), carbohydrate (structure carbohydrate (SC) and non-structure carbohydrate (NSC)), and environmental variables in Pinus thunbergii plantations from coast to inland, with wind speed decreasing. As expected, wind, accounting for 19–69% of the total variation, was the most dominant environmental variable determining the leaf traits. Y(II) and NSC increased, while SC and SC/NSC decreased along the coast-inland gradients (p < 0.01). These results confirmed that, although carbohydrate production decreased, SC allocation increased with increasing wind load. SLA and needle thickness decreased, while cuticle thickness and epidermal thickness increased from coast to inland. Needle thickness and cuticle thickness showed strong correlations to SC/NSC. These variations indicated that carbohydrate categories allocation related to variations of needle morphology and anatomy for P. thunbergii under wind, because of more SC allocation in leaf to support tensile strength and hardness of the cell wall under wind. Therefore, allocation between SC and NSC may be helpful for understanding the long-term adaptation of plants to wind load.

Detecting carbohydrate molecular structural makeup in different types of cereal grains and different cultivars within each type of grain grown in semi-arid area using FTIR spectroscopy with uni- and multi-variate molecular spectral analyses

Various cereal grains including barley (three varieties: moderate (MT), cold (CT), and dry (DT)), corn (two hybrids: early (EM) and late maturity (LM)), and sorghum (two varieties: new type (NT) and conventional type (OT)) were used to characterize carbohydrate molecular structural features in cereal grains using Fourier transform infrared spectroscopy (FTIR). Molecular spectral peak bands assessed included: A_860 (non structure carbohydrate: ca. 880–818cm−1), A_928 (non-structure carbohydrate: ca. 946–881cm−1), total carbohydrate (TCHO: ca.1186–947cm−1) with three major component peaks (ca. 1017, 1074, and 1152cm−1), and cellulosic compounds (ca. 1291–1186cm−1). The results showed carbohydrate molecular spectral features differed among cereal grains. The infrared absorbance intensities of A_860, A_928, TCHO component peaks, and cellulosic compounds were greater for barley than those for both corn and sorghum grains which were similar to each other. These spectral intensities were greater for EM than those for LM in corn hybrids, and greater for OT than for NT in sorghum cultivars. These structural differences may impact the carbohydrate utilization and availability of cereal grains. Results from multivariate spectral analysis indicated that there were still structural similarities in cereal grains and varieties within each type of grain.

Synthesis of new enantiopure poly(hydroxy)aminooxepanes as building blocks for multivalent carbohydrate mimetics.

New compounds with carbohydrate-similar structure (carbohydrate mimetics) are presented in this article. Starting from enantiopure nitrones and lithiated TMSE-allene we prepared three 1,2-oxazine derivatives which underwent a highly stereoselective Lewis acid-induced rearrangement to give bicyclic products in good yield. Subsequent reductive transformations delivered a library of new poly(hydroxy)aminooxepane derivatives. The crucial final palladium-catalyzed hydrogenolysis of the 1,2-oxazine moiety was optimized resulting in a reasonably efficient approach to a series of new seven-membered carbohydrate mimetics.

Food Technological Applications for Optimal Nutrition: An Overview of Opportunities for the Food Industry

Abstract: An increasingly important determinant in food choice is the growing consumer concern about nutrition and health. This focusing of consumer interest on the food supply, and also extensive research and technological developments in food science will provide further opportunities for new product development. The Food‐Based Dietary Guidelines of the World Health Organization (WHO) and European Union (EU) legislation on health claims play an important role in regulating information to the public about a wholesome diet and for improving the availability and affordability of nutritious food choices to consumers. More specifically, the food industry can contribute by reducing the number of energy‐dense products; by improving the nutrient profile of processed food through the reduction of salt, added sugar, trans‐fatty acid, and saturated fat content. As a result, food science and technology are prompted to create a new framework for these food‐based dietary guidelines, principally in the areas of food physics, methods of food storage and preservation, nutrient restoration and fortification of foods, and the development of health‐focused designer foods and functional foods. The aim of this review is to provide an overview of some further opportunities for new product development and nutrition research. Some topics related to the energy reduction of foods include: dilution and structure design, carbohydrate and/or fat substitutes, and inhibition of enzymes in carbohydrate and/or fat digestion; additionally, regulation of some metabolic functions with food‐derived bioactive peptides and probiotics, and enrichment of foods with bioactive compounds are reviewed in this overview as the most promising issues.

Novel Fold and Carbohydrate Specificity of the Potent Anti-HIV Cyanobacterial Lectin from Oscillatoria agardhii

Oscillatoria agardhii agglutinin (OAA) is a recently discovered cyanobacterial lectin that exhibits potent anti-HIV activity. Up to now, only its primary structure and carbohydrate binding data have been available. To elucidate the structural basis for the antiviral mechanism of OAA, we determined the structure of this lectin by x-ray crystallography at 1.2 Å resolution and mapped the specific carbohydrate recognition sites of OAA by NMR spectroscopy. The overall architecture of OAA comprises 10 β-strands that fold into a single, compact, β-barrel-like domain, creating a unique topology compared with all known protein structures in the Protein Data Bank. OAA sugar binding was tested against Man-9 and various disaccharide components of Man-9. Two symmetric carbohydrate-binding sites were located on the protein, and a preference for Manα(1-6)Man-linked sugars was found. Altogether, our structural results explain the antiviral activity OAA and add to the growing body of knowledge about antiviral lectins.

Purification, crystallization and preliminary X-ray study of the fungal laccase from Cerrena maxima.

Laccases are members of the blue multi-copper oxidase family that oxidize substrate molecules by accepting electrons at a mononuclear copper centre and transferring them to a trinuclear centre. Dioxygen binds to the trinuclear centre and, following the transfer of four electrons, is reduced to two molecules of water. Crystals of the laccase from Cerrena maxima have been obtained and X-ray data were collected to 1.9 A resolution using synchrotron radiation. A preliminary analysis shows that the enzyme has the typical laccase structure and several carbohydrate sites have been identified. The carbohydrate chains appear to be involved in stabilization of the intermolecular contacts in the crystal structure, thus promoting the formation of well ordered crystals of the enzyme. Here, the results of an X-ray crystallographic study on the laccase from the fungus Cerrena maxima are reported. Crystals that diffract well to a resolution of at least 1.9 A (R factor = 18.953%; R(free) = 23.835; r.m.s.d. bond lengths, 0.06 A; r.m.s.d. bond angles, 1.07 degrees) have been obtained despite the presence of glycan moieties. The overall spatial organization of C. maxima laccase and the structure of its copper-containing active centre have been determined by the molecular-replacement method using the laccase from Trametes versicolor (Piontek et al., 2002) as a structural template. In addition, four glycan-binding sites were identified and the 1.9 A X-ray data were used to determine the previously unknown primary structure of this protein. The identity (calculated from sequence alignment) between the C. maxima laccase and the T. versicolor laccase is about 87%. Tyr196 and Tyr372 show significant extra density at the ortho positions and this has been interpreted in terms of NO(2) substituents.

Primary structure and unusual carbohydrate moiety of functional unit 2- c of keyhole limpet hemocyanin (KLH)

The complete amino acid sequence of the Megathura crenulata hemocyanin functional unit KLH2- c was determined by direct sequencing and matrix-assisted laser desorption ionization mass spectrometry of the protein, and of peptides obtained by cleavage with EndoLysC proteinase, chymotrypsin and cyanogen bromide. This is the first complete primary structure of a functional unit c from a gastropod hemocyanin. KLH2- c consists of 420 amino acid residues. Circular dichroism spectra indicated approx. 31% β-sheet and 29% α-helix contents. A multiple sequence alignment with other molluscan hemocyanin functional units revealed average identities between 41 and 49%, but 55% in case of Octopus hemocyanin functional unit c which is the structural equivalent to KLH2- c. KLH2- c has a molecular mass of approx. 48 kDa as calculated from its sequence and a measured mass of approx. 56 kDa; the mass difference is attributed to the sugar side chains usually decorating molluscan hemocyanin. However, inspection of the sequence of KLH2- c revealed no potential N-linked carbohydrate attachment sites, and this was supported by its inability to bind concanavalin A. Also KLH1- c was unreactive, whereas most, if not all, other functional units of KLH1 and KLH2 reacted positively to this lectin. On the other hand, peanut agglutinin specifically binds KLH2- c, indicating the presence of O-glycosidically linked carbohydrates in this functional unit. This contrasts to all other KLH functional units (including KLH1- c), which lack O-linked glycosides. The present results are discussed in view of the recent X-ray structure of the functional unit g from Octopus hemocyanin, and a published record of the Thomsen Friedenreich tumor antigenic epitope in KLH.

Dynamics of the anaerobic utilization of organic substrates in an anaerobic/aerobic sequencing batch reactor

The operation of an anaerobiclaerobic Sequencing Batch Reactor (SBR) with synthetic feed (glucose as the sole organic substrate) demonstrated periods of non-EBPR and EBPR operation to the absence of polyhydroxy alkanoate (PHA) storage. The glucose added as feed disappeared rapidly (within 10 minutes) and none of the normal fermentation products were detected in the supernatant in either mode of operation. Anaerobic/aerobic batch trials using the biomass from the SBR were conducted to examine the processes taking place and a rapid uptake of glucose was confirmed and was accompanied by a fall m pH. The biomass also demonstrated that anaerobic PHB storage occurred when acetate feed was used. The biomass was characterized by a large population of "G" bacteria, the absence of filamentous microorganisms, compact floc structure and high total carbohydrate (ca 40% w/w). The supernatant was characterized by low COD, TOC and the virtual absence of the glucose substrate. The observations indicate that EBPR was possible under circumstances which are not consistent with accepted mechanisms.

Dynamics of the anaerobic utilization of organic substrates in an anaerobic/aerobic sequencing batch reactor

The operation of an anaerobic/aerobic Sequencing Batch Reactor (SBR) with synthetic feed (glucose as the sole organic substrate) demonstrated periods of non-EBPR and EBPR operation in the absence of polyhydroxy alkanoate (PHA) storage. The glucose added as feed disappeared rapidly (within 10 minutes) and none of the normal fermentation products were detected in the supernatant in either mode of operation. Anaerobic/aerobic batch trials using the biomass from the SBR were conducted to examine the processes taking place and a rapid uptake of glucose was confirmed and was accompanied by a fall in pH. The biomass also demonstrated that anaerobic PHB storage occurred when acetate feed was used. The biomass was characterized by a large population of “G” bacteria, the absence of filamentous microorganisms, compact floc structure and high total carbohydrate (ca 40% w/w). The supernatant was characterized by low COD, TOC and the virtual absence of the glucose substrate. The observations indicate that EBPR was possible under circumstances which are not consistent with accepted mechanisms.

Wheat germ agglutinin inhibits the C5a receptor interaction: implications for receptor microheterogeneity and ligand binding site.

Wheat germ agglutinin (WGA) has been shown to inhibit the interaction of C5a with the C5a receptor on both polymorphonuclear neutrophils (PMNs) and the histiocytic cell line U937. The level of inhibition with isolated receptor preparations is 100%, and on intact cells 10 to 20% of the receptor population appear to retain their ability to bind C5a in the presence of WGA. In contrast, this lectin completely inhibits the C5a-mediated degranulation of PMN primary and secondary granules, suggesting that the population of C5a receptors responsible for mediating degranulation is also recognized by WGA. More than 50% of the receptors appear to be blocked before an effect on degranulation occurs. This inhibition by WGA does not appear to be due to down-regulation of C5a receptors from the cell surface, excessive aggregation of receptor sites, or interaction of WGA with the carbohydrate portion of the C5a molecule. The inhibition is reversed by N-acetylglucosamine but not by sialic acid. This effect appears to be specific for WGA because various other lectins do not inhibit the C5a receptor interaction. That the inhibition by WGA is due to direct binding of the lectin to N-acetylglucosamine residues on the C5a receptor is strongly supported by the ability of the cross-linked C5a-receptor complex to bind to and be specifically eluted from a WGA-Affigel affinity matrix. These observations are consistent with hypothesis that the population of C5a receptors on leukocytes exhibits microheterogeneity with respect to structure (carbohydrate content) and/or function.

Changes in Morphological Stage of Development, Canopy Structure, and Root Nonstructural Carbohydrate Reserves of Alfalfa Following Control of Potato Leafhopper (Homoptera: Cicadellidae) and Weed Populations

Impacts of weed and insect control measures on populations of the potato leafhopper, Empoasca fabae (Harris), and various alfalfa plant parameters were studied from 1983 through 1985. Application of summer annual herbicides for the control of grass weeds produced a dual effect, increasing alfalfa density by reducing competition from grass weeds, and encouraging the presence of potato leafhopper (PLH) and thus increased crop damage by reducing plant stage of growth and stem length. Use of winter annual herbicides was associated with an increase in root total nonstructural carbohydrates (TNC) and reduced stem density, suggesting possible herbicide damage to the alfalfa crop. Insecticide applications reduced PLH damage (hopperburn) and were associated with a more fully developed and taller alfalfa crop. By separately correlating both PLH population levels and incidence of hopperburn with the plant parameters analyzed, identical trends were noted. Thus, hopperburn may be a possible indicator of insect abundance.