Search

A Chlorella sp. was isolated from a brackish water high-altitude lake of the Northern-Western Himalayas and selected on the basis of its ability to secrete substantial amount of exopolysaccharide (EPS). The purified EPS was estimated to have molecular weight (Mw) of approximately 1.52 × 105 Da with polydispersity of 2.315 and showed amorphous dense morphology. The production of the EPS was maximized to ≅ 460 mg L−1 through nutrient optimisation. The monosaccharide composition of EPS showed the presence of hexose (glucose, galactose mannose), pentose (xylose, arabinose), deoxy (rhamnose), amino (galactosamine) and acid (galacturonic acid and glucuronic) sugars. The one-dimensional proton NMR and FT-IR analysis confirmed the presence of aliphatic groups. The pseudo-plastic rheology and the antioxidant activity of the EPS suggested its suitability for industrial applications. Importantly, the isolated polysaccharide exhibits its potential for easy and single pot biosynthesis of polysaccharide-capped gold-nanoparticles (AuNPs). The polysaccharide-capped AuNPs displayed exceptional stability at extended pH range and high salinity and suggest that this algal EPS can be utilized for its biotechnological application.

Human milk oligosaccharides (HMOs) have attracted much attention in recent years not only as a prebiotic factor but also in particular as an essential component of infant nutrition in relation to their impact on innate immunity. The backbone structures of complex HMOs generally contain single or repetitive lacto-N-biose (type 1) or lactosamine (type 2) units in either linear or branched chains extending from a lactose core. While all known branched structures originate from the 3,6-substitution of the lactosyl core galactose, we here describe a new class of HMOs that tentatively branch at the terminal galactose of 6'-galactosyllactose. Another novel feature of this class of HMOs was found in linear oligo-galactosyl chains linked to one of the N-acetylglucosamine (GlcNAc) branches. The novel structures exhibit general formulas with hexose versus hexosamine contents of 5/2 to 8/2 and can be designated as high-galactose (HG)-HMOs. In addition, up to three fucosyl residues are linked to the octa- to dodecasaccharides, which were detected in two human milk samples from the Lewis blood-group-defined donors. Structural analyses of methylated glycans and their alditols comprised matrix-assisted laser desorption ionization mass spectrometry, electrospray-(collision-induced dissociation) mass spectrometry and linkage analyses by gas chromatography-mass spectrometry of the derived partially methylated alditol acetates. Enzymatic degradation by the application of β1-3,4-specific galactosidase supported the presence of terminal galactose-linked β1-6 to one of the two GlcNAc branches. The mass spectrometry glycomic data have been deposited at the GlycoPOST archive with the data set identifier GPST000191 (Username: franz.hanisch@uni-koeln.de; Password: Soma1Dita2Carb. Watanabe, Y. GlycoPOST realizes FAIR principles for glycomics mass spectrometry data. Nucleic Acids Res. 2021, 49, D1523-D1528).

UDP-glucose 4-epimerase (GalE) catalyzes the interconversion of UDP-glucose (UDP-Glc) and UDP-galactose (UDP-Gal) and/or the interconversion of UDP-N-acetylglucosamine (UDP-GlcNAc) and UDP-N-acetylgalactosamine (UDP-GalNAc) in sugar metabolism. GalEs belong to the short-chain dehydrogenase/reductase superfamily, use a conserved 'transient keto intermediate' mechanism and have variable substrate specificity. GalEs have been classified into three groups based on substrate specificity: group 1 prefers UDP-Glc/Gal, group 3 prefers UDP-GlcNAc/GalNAc, and group 2 has comparable activities for both types of the substrates. The phylogenetic relationship and structural basis for the specificities of GalEs revealed possible molecular evolution of UDP-hexose 4-epimerases in various organisms. Based on the recent advances in studies on GalEs and related enzymes, an updated view of their evolutional diversification is presented.

Cancer cells increase their metabolic activity by enhancing glucose uptake through overexpression of hexose transporters (Gluts). Gluts also have the capacity to transport other molecules besides glucose, including fructose, mannose, and dehydroascorbic acid (DHA), the oxidized form of vitamin C. The majority of research studies in this field have focused on the role of glucose transport and metabolism in cancer, leaving a substantial gap in our knowledge of the contribution of other hexoses and DHA in cancer biology. Here, we summarize the most recent advances in understanding the role that the multifunctional transport capacity of Gluts plays in biological and clinical aspects of cancer, and how these characteristics can be exploited in the search for novel diagnostic and therapeutic strategies.

Pentose and hexose sugars are abundant constituents of waste biomass, making them sustainable, chiral building blocks for organic synthesis. The demand for chiral saturated heterocyclic rings from the pharmaceutical industry is increasing as they provide well-defined three-dimensional frameworks that show increased metabolic resistance. Through the formation of thioacetals, sugars may be manipulated in their straight-chain form and dehydrated selectively under basic conditions at C-2. This approach was applied to an array of sugars and extended to the production of useful chiral THFs via further selective dehydration reactions.

Background Increased soil salt concentration decreases productivity and changes the physiological and chemical properties of plants. Various omics technologies have been used to understand the salt response in plants but overall changes in the metabolite profiles of spinach (Spinacia oleracea L.) under salt stress have not been studied. In this article, therefore, the changes in mineral and metabolite profiles of spinach plants cultivated with different NaCl concentrations of 0-200 mmol L-1 in the irrigation water were analyzed to investigate the effect of salt stress on nutritional quality. Results Increasing NaCl concentration decreased plant growth due to mineral imbalance. The amounts of minerals (K+ , Ca2+ , and Fe2+ ) were reduced with increasing NaCl concentration, resulting in altered ratios of Na+ :K+ and Na+ :Ca2+ . The change in the mineral ratios due to NaCl irrigation led to a decrease in the height and an increase in the weight of spinach. Moreover, the profiles of 32 metabolites, including flavonoids, amino acids, acidic compounds, sugars, and lipid-related compounds, were altered by NaCl irrigation; most of them showed decreased levels. In particular, at 200 mmol L-1 NaCl, the levels of sucrose, glutamic acid, hexose sugars, and acidic compounds significantly decreased upon NaCl irrigation. Based on these metabolites, a salt-stress-related spinach metabolomic pathway was proposed. Conclusion Sodium chloride irrigation increased mineral imbalance, resulting in decreased plant growth, and the levels of most metabolites involved in energy production, sensory quality, and health benefits decreased with NaCl irrigation. The results suggest that NaCl irrigation negatively affects the nutritional quality of spinach. © 2020 Society of Chemical Industry.

Bacterial nonulosonic acids such as pseudaminic acids and others constitute a family of 9-carbon monosaccharides that contain a common 3-deoxy-2-ketoacid fragment but differ in their stereochemistries at 5 stereogenic centers between C-4 to C-8. Their unique structures make them attractive targets for use as antigens in vaccinations to combat drug-resistant bacterial infections and their challenging stereochemistries have attracted considerable attention from chemists. In this work we report the development of an improved synthesis for 2,4-di-N-acetyl-L-altrose (L-2,4-Alt-diNAc), which is a key hexose required for the chemical and chemoenzymatic synthesis of pseudaminic acids. Using L-fucose as a starting material, our synthesis overcomes several pitfalls in previously reported syntheses.

A reduction in cytosolic invertase in Arabidopsis roots changes the sucrose to hexose ratio, resulting in profound alterations of metabolism, growth, development, and patterns of gene expression., Plant roots depend on sucrose imported from leaves as the substrate for metabolism and growth. Sucrose and hexoses derived from it are also signalling molecules that modulate growth and development, but the importance for signalling of endogenous changes in sugar levels is poorly understood. We report that reduced activity of cytosolic invertase, which converts sucrose to hexoses, leads to pronounced metabolic, growth, and developmental defects in roots of Arabidopsis (Arabidopsis thaliana) seedlings. In addition to altered sugar and downstream metabolite levels, roots of cinv1 cinv2 mutants have reduced elongation rates, cell and meristem size, abnormal meristematic cell division patterns, and altered expression of thousands of genes of diverse functions. Provision of exogenous glucose to mutant roots repairs relatively few of the defects. The extensive transcriptional differences between mutant and wild-type roots have hallmarks of both high sucrose and low hexose signalling. We conclude that the mutant phenotype reflects both low carbon availability for metabolism and growth and complex sugar signals derived from elevated sucrose and depressed hexose levels in the cytosol of mutant roots. Such reciprocal changes in endogenous sucrose and hexose levels potentially provide rich information about sugar status that translates into flexible adjustments of growth and development.

Aim: the present study was carried out to compare the glycoproteins in normal individuals and in patients with hypothyroidism and hyperthyroidism. Materials and Methods: This study was carried out at Meenakshi Medical College & Research Institute on total 90 individuals, 30 hypothyroid patients, 30 hyperthyroid patients and 30 normal individuals were taken as controls. Each group consisted of both sexes and the participants were between the ages of 25 to 50 years. Various parameters like protein bound fucose, hexose, hexosamine, sialic acid and thyroid function test which include ft3, ft4 and TSH were measured in all the 90 individuals. The data was collected and analyzed. Result: Potein bound hexose, sialic acid, fucose and hexosamine decreased in hypothyroid patients when compared to normal individuals, whereas in patients with hyperthyroidism only protein bound hexose and fucose levels increased significantly whereas sialic acid, hexose were decreased when compared to normal individuals. Conclusion: Serum Glycoproteins can be additional markers to ft3, ft4 and TSH and aid in diagnosis and management of thyroid disorders. Keywords: Glycoproteins, Thyroid function test, Hyperthyroidism, Hypothyroidism.

Two new phenolic glucosides, including a new O-glycoside (1) and a new C-glycoside (2), were isolated from a marine-derived fungus Aspergillus sp. The structures of new compounds were elucidated through interpretations of spectroscopic evidence and high-resolution electrospray ionization mass spectrometry. The hexose unit of 1 was identified as β-D-glucose by comparison with an authentic sample via HPLC after acid hydrolysis and derivatization. All compounds were evaluated for their ability to inhibit LPS-induced NO production in RAW264.7 macrophages, but none of them displayed significant activity.

Chinese cabbage (Brassica campestris L. ssp. pekinensis) is a widely cultivated vegetable crop in eastern Asia. The flavor of Chinese cabbage was mostly affected by soluble sugar content. To make clear its accumulation and regulation mechanism, the content of different sugars in the internal blade (IB), the internal midrib (IM), the external blade (EB) and the external midrib (EM) of the leafy head were determined during the leafy head formation. The results showed that fructose was the major sugar accumulated in the internal tissues, followed by glucose. IM was the main tissue of sugar accumulation with the highest contents of total soluble sugar and hexose at harvest. RNA sequencing data of the four tissues at 90 DAS showed that the number of differentially expressed genes (DEGs) in IB and IM was the least (2408), while the number of DEGs between the external and internal tissues ranged from 6037 to 8674. The enzyme genes differentially expressed in ‘starch and sucrose metabolism’ pathway was detected during the leafy head development. The expression of two sucrose synthase (SUS) genes, SUS1a (Bra002332) and SUS1b (Bra006587), were always higher in IM than in the other tissues, and positively correlated with the content of total soluble sugar and hexose, respectively. SUS1 was considered to play an important role in leafy head development and sugar accumulation in Chinese cabbage.

—The investigation of the mechanisms of cytosolic protein degradation is of great fundamental and applied importance. The decrease in the specific activity of formaldehyde dehydrogenase (Fldh1) and formate dehydrogenase (Fdh1) in the wild type strain GS200, the strain with the deletion of the GSS1 hexose sensor gene, and the strain that is defective in autophagy pathway SMD1163 of K. phaffii during short-term and long-term induction with methanol, with or without the addition of the MG132 (proteasome degradation inhibitor), was investigated. It was shown that the duration of cell incubation on methanol had no particular effect on the inactivation of enzymes. The effect of the proteasome inhibitor MG132 was insignificant. Catabolic inactivation of cytosolic and peroxisomal enzymes was damaged in the gss1Δ mutant since glucose signaling was impaired. Fldh1 and Fdh1 are probably degraded via the vacuolar pathway regardless of the duration of methanol induction.

Water stress is a primary trigger for reducing grain number per spike in wheat during the reproductive period. However, under stress conditions, the responses of plant organs and the interactions between them at the molecular and physiological levels remain unclear. In this study, when water stress occurred at the young microspore stage, RNA-seq data indicated that the spike had 970 differentially expressed genes, while the stem, comprising the two internodes below the spike (TIS), had 382. Abscisic acid (ABA) signal transduction genes were down-regulated by water stress in both these tissues, although to a greater extent in the TIS than in the spike. A reduction in sucrose was observed, and was accompanied by increases in cell wall invertase (CWIN) and sucrose:sucrose 1-fructosyl-transferase (1-SST) activities. Hexose and fructan were increased in the TIS but decreased in the spike. ABA was increased in the spike and TIS, and showed significant positive correlation with CWIN and 1-SST activities in the TIS. Overall, our results suggest that water stress induces the conversion of sucrose to hexose by CWIN, and to fructan by 1-SST, due to increased down-regulation of ABA signal transduction related-genes in the TIS; this leads to deficient sucrose supply to the spike and a decrease in grain number., Deficient sucrose and weak expression of ABA signal transduction genes in the spike may be the major reasons for the decrease in grain number under water stress.

Stereoisomeric hexoses are present in almost all biological organisms in the forms of aldoses and ketoses, with diverse physiological and pathophysiological functions. Accurate and simultaneous quantification is vital for understanding their functions individually. However, such analysis remains challenging owing to their highly similar behavior in chromatography and mass spectrometry. By combining the pre-column 3-nitrophenylhydrazine derivatization and ultrahigh performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS), here, we developed a method for simultaneous quantification of five important stereoisomeric hexoses including D-glucose, D-galactose, D-mannose, D-fructose and L-sorbose representing both aldoses and ketoses. The method achieved baseline-separation for all these five derivatized hexoses chromatographically and had high sensitivity (LOD, femtomole on column), excellent linearity (R2 > 0.995) and efficiency with stable-isotope dilution. With this method, we further quantified these hexoses in nine biological matrices including human biofluids (serum, urine and saliva), human cells, human and mouse feces, rat liver tissue, mung-bean seeds and peach pulp. The results provided quantitative data for these hexoses in multiple biological samples and showed significant concentration diversity for these hexoses in different biological samples, which demonstrated the applicability of the method for simultaneous quantification of these hexose phenotypes of biological systems. • An optimized UHPLC-MS/MS method based on 3-nitrophenylhydrazine derivatization. • Simultaneous quantification of five stereoisomeric hexoses. • Quantitative results for these five hexoses in nine different biological matrices.

Biomass, a type of inexpensive and sustainable carbonaceous resource, is considered a promising substitute for fossil fuel-derived products in various fields. Hexose and pentose sugars are the buil...

The aim of this work was to evaluate the general role of the vacuolar pyrophosphatase proton pump (V-PPase) in sucrose accumulation in citrus species. First, three citrus V-PPase genes, designated CsVPP-1, CsVPP-2, and CsVPP-4, were identified in the citrus genome. CsVPP-1 and CsVPP-2 belonging to citrus type I V-PPase genes are targeted to the tonoplast, and CsVPP-4 belonging to citrus type II V-PPase genes is located in the Golgi bodies. Moreover, there was a significantly positive correlation between transcript levels of type I V-PPase genes and sucrose, rather than hexose, content in fruits of seven citrus cultivars. Drought and abscisic acid treatments significantly induced the CsVPP-1 and CsVPP-2 transcript levels, as well as the sucrose content. The overexpression of type I V-PPase genes significantly increased PPase activity, decreased pyrophosphate contents, and increased sucrose contents, whereas V-PPase inhibition produced the opposite effect in both citrus fruits and leaves. Furthermore, altering the expression levels of type I V-PPase genes significantly influenced the transcript levels of sucrose transporter genes. Taken together, this study demonstrated that CsVPP-1 and CsVPP-2 play key roles in sucrose storage in the vacuole by regulating pyrophosphate homeostasis, ultimately the sucrose biosynthesis and transcript levels of sucrose transport genes, providing a novel lead for engineering or breeding modified taste in citrus and other fruits.

To clarify the complex regulatory relationship between changes in sugar content and leaf senescence during the grain-filling stage of rice, genotype-dependent differences in sugar content and the temporal transcriptional patterns of genes involved in sugar signaling pathways were determined in mutant rice exhibiting early leaf senescence and its wild type Zhefu 142. The effects of exogenous glucose or sucrose on the senescence of detached leaves under dark conditions were also investigated. Chlorophyll, soluble sugar, sucrose and fructose contents decreased, whereas electrolytic leakage and malondialdehyde levels increased in mutant leaves at the grain-filling stage. These results suggested that sugar starvation is positively correlated with the early leaf senescence of mutant plants. Detached leaf segments incubated in exogenous sugar solutions under dark conditions exhibited delayed senescence. The high expression of Hxk1 in leaves of mutant plants at the initial grain-filling stage suggested that Hxk1 is involved in the hexose-sensing process at the early stage of leaf senescence. The low expression levels of Hxk2 and Frk1 in the senescing leaves of mutant rice during the grain-filling stage are indicative of weakened hexose phosphorylation. In addition, the high expression levels of SuSy1, SuSy2 and SuSy4 in leaves of mutant plants at the initial grain-filling stage are accompanied by the high transcript levels of SUT1, which favor sucrose translocation and remobilization from the early senescing leaves of mutant rice. The relatively reduced transcript levels of chFBP, cyFBP, SPS1, SPS2 and SPS6 indicated that during the grain-filling stage, sucrose biosynthesis is weakened in the senescing leaves of mutant rice. Keywords: rice, sugar starvation, leaf senescence, signaling pathway, detached leaf segment, grain-filling stage

Shading and drought stresses during the flowering stage of maize (Zea mays L.) production cause seed losses, making them two of the major limitations to yield output. The mechanisms by which the two stresses reduce seed numbers are still unclear. Here, different durations of shading and different degrees of drought were applied, accompanied by synchronous pollination, and the early development of fertilized apical kernels was suppressed, with decreases in sucrose and hexose levels and the promotion of ethylene emission. Consequently, the two distinct stresses led to a common consequence: up to 20–30% of the fertilized kernels were aborted at maturity, which accounted for the yield losses under both scenarios. Notably, by removing the shading net or the preventing pollination of the basal kernels, the apical kernels, were partial or fully restored from abortion, with an increased sugar status and suppressed ethylene emission, even under stress conditions. We found that dry matter accumulation within the early stage was positively correlated with sucrose and hexose contents but negatively correlated with ethylene emission. By altering the sugar status within developing kernels, we verified the antagonistic relationships between endogenous ethylene emission and soluble sugars, including sucrose and hexoses. Collectively, these findings suggest that the equilibrium between sugars and ethylene may determine the development or abortion of early developing kernels exposed to environmental stresses.

The acidic catalysts play pivotal roles in the dehydration of hexose into 5-hydroxymethylfurfural (HMF). However, these acidic catalysts were usually exogenously added or self-made, which increase ...

Ferulate 5-hydroxylase (F5H) is a limiting enzyme involved in biosynthesizing sinapyl (S) monolignol in angiosperms. Genetic regulation of F5H can influence S monolignol synthesis and therefore improve saccharification efficiency and biofuel production. To date, little is known about whether F5H is post-transcriptionally regulated by endogenous microRNAs (miRNAs) in woody plants. Here, we report that a microRNA, miR6443, specifically regulates S lignin biosynthesis during stem development in Populus tomentosa. In situ hybridization showed that miR6443 is preferentially expressed in vascular tissues. We further identified that F5H2 is the direct target of miR6443. Overexpression of miR6443 decreased the transcript level of F5H2 in transgenic plants, resulting in a significant reduction in S lignin content. Conversely, reduced miR6443 expression by short tandem target mimics (STTM) elevated F5H2 transcripts, therefore increasing S lignin composition. Introduction of a miR6443-resistant form of F5H2 into miR6443-overexpression plants restored lignin ectopic composition, supporting that miR6443 specifically regulated S lignin biosynthesis by repressing F5H2 in P. tomentosa. Furthermore, saccharification assays revealed decreased hexose yields by 7.5-24.5% in miR6443-overexpression plants compared with the wild-type control, and increased hexoses yields by 13.2-14.6% in STTM6443-overexpression plants. Collectively, we demonstrate that miR6443 modulates S lignin biosynthesis by specially regulating F5H2 in P. tomentosa.

Pollen tubes require a high amount of sugars to sustain high growth rate. Sucrose, the main transport form of sugars, can serve as energy supply and as signaling molecules for pollen tube growth. We report the functional characterization of CsHT11, which is a sugar transporter protein/hexose transport protein (STP/HT). CsHT11 shares high homology with the characterized cucumber hexose transporter CsHT1, a pollen-specific gene. Analysis of CsHT11 mRNA and CsHT11 promoter-reporter gene studies revealed CsHT11 specifically expressed in pollen and pollen tubes. Subcellular localization indicated that CsHT11 is a plasma membrane transporter. Heterologous expression in yeast suggests that CsHT11 is an energy-dependent hexose/H+ symporter, with a wide variety of substrate specificity, including glucose, fructose, galactose, and mannose. In vitro pollen germination of different sugars shows that the expression of CsHT11 is significantly increased with higher sucrose content, but not with higher glucose or fructose content, thereby implying that CsHT11 may be involved in sucrose signal transduction. Thus, CsHT11 might have an essential effect on pollen development and pollen tube growth in cucumber.

Late oxidation of hexose based building blocks or the use of uronic acid containing building blocks are two complementary strategies in the synthesis of glycosaminoglycans, the latter simplifiying the later stages of the process. Here we report the synthesis and evaluation of various disaccharide donors-uronic acids and their pyranose equivalents-for the synthesis of heparan sulfate, using an established protective group strategy. Hexose based "imidate" type donors perform well in the studied glycosylations, while their corresponding uronate esters fall short; a uronate ester thioglycoside performs equal to, if not better than, a hexose thioglycoside equivalent.

Rapid pollen tube growth requires uptake of Suc or its hydrolytic products, hexoses, from the apoplast of surrounding tissues in the style. Due to species-specific sugar requirements, reliance of pollen germination and tube growth on cell wall invertase and Suc or hexose transporters varies between species, but it is not known if plants have a sugar transporter that mediates the uptake of both hexose and Suc for pollen tube growth. Here, we show that a sugar transporter protein in apple (Malus domestica), MdSTP13a, takes up both hexose and Suc when expressed in yeast, and is essential for pollen tube growth on Glc and Suc but not on maltose. MdSTP13a-mediated direct uptake of Suc is primarily responsible for apple pollen tube growth on Suc medium. Sorbitol, a major photosynthate and transport carbohydrate in apple, modulates pollen tube growth via the MYB transcription factor MdMYB39L, which binds to the promoter of MdSTP13a to activate its expression. Antisense repression of MdSTP13a blocks sorbitol-modulated pollen tube growth. These findings demonstrate that MdSTP13a takes up both hexose and Suc for sorbitol-modulated pollen tube growth in apple, revealing a situation where acquisition of sugars for pollen tube growth is regulated by a sugar alcohol.

Nostoc microscopicum an effective extracellular polymers (EPs) synthesizer among cyanobacteria was isolated, and its elementary morphological features were defined with the aid of light microscope and CLSM. Bioseparation of EPs from 44 days-aged culture gave 0.90 g/L of the dry powdered extract. Chemical quantification of EPs showed the presence of 550 mg/g of carbohydrate and 395 mg/g of protein. HPLC results depicted the presence of mannose (monomer sugar), fucose (hexose deoxy sugar), mannitol (sugar alcohol) and N- acetylglucosamine (glycoprotein) in the EPs. The vibration-based spectrum produced by FT-IR proves the β-Sheet structure of EPs glycoprotein and the size as 45 kDa by performing SDS-PAGE. Bactericidal activity evaluation of EPs on Pseudomonas aeruginosa and Staphylococcus aureus co-expressed the MIC value as 125 μg/mL, while zone of inhibition was 12 mm for Gram-negative and 8 mm for Gram-positive bacteria. Biofilm inhibition assay was effective in 1.0 mg/mL concentration of EPs in both bacterial strains with a mean rate of 60 percentages which was further confirmed using confocal laser scanning microscopic imaging. This natural polymeric extract of Nostoc microscopicum indicates its possible applications in bactericidal and biofilm inhibition.

Hexose transporters play many important roles in plant development. However, the role of hexose transporter in secondary cell wall growth has not been reported before. Here, we report that the hexose transporter gene CsHT3 is mainly expressed in cells with secondary cell walls in cucumber. Spatiotemporal expression analysis revealed that the transcript of CsHT3 mainly accumulates in the stem, petiole, tendril, and peduncle, all of which contain high cellulose levels. Immunolocalization results show that CsHT3 is localized at the sclereids in young peduncles, shifts to the phloem fiber cells during peduncle development, and then shifts again to the companion cells when the development of secondary cell walls is almost completed. Carboxyfluoresce unloading experiment indicated that carbohydrate unloading in the phloem follows an apoplastic pathway. Overexpression of CsHT3 in cucumber plant can improve the cellulose content and cell wall thickness of phloem fiber cells in the peduncle. The expression of cellulose synthase genes were increased in the CsHT3 overexpression plants. These results indicated that CsHT3 may play an important role in cellulose synthesis through promoting the expression of cellulose synthase genes.

The degradative condensation of hexose, which originates from the C-C cleavage of hexose and condensation of degraded hexose fragment, is one of the possible reaction pathways for the formation of humins in hexose dehydration to 5-hydroxymethylfurfural (HMF). Herein, the impacts of several polar aprotic solvents on the degradative condensation of fructose to small-molecule carboxylic acids and oligomers (possible precursors of humins) are reported. In particular, a close relationship between the tautomeric distribution of fructose in solvents and the mechanism of degradative condensation is demonstrated. Typically, α-fructofuranose in 1,4-dioxane and acyclic open-chain fructose in THF favor the conversion of fructose to formic acid and oligomers; α-fructopyranose in γ-valerolactone or N-methylpyrrolidone favors levulinic acid and oligomers, whereas β-fructopyranose in 4-methyl-2-pentanone favors acetic acid and corresponding oligomers. This close correlation highlights a general understanding of the solvent-controlled formation of oligomers, which represents an important step toward the rational design of effective solvent systems for HMF production.

Furfural is a common furan inhibitor formed due to dehydration of pentose sugars, like xylose, and acts as an inhibitor of microbial metabolism. Overexpression of NADH-specific FucO and deletion of NADPH-specific YqhD had been a successful strategy in the past in conferring tolerance against furfural in Escherichia coli, which highlights the importance of oxidoreductases in conferring tolerance against furfural. In a screen consisting of various oxidoreductases, dehydrogenases, and reductases, we identified the yghA gene as an overexpression target to confer tolerance against furfural. YghA preferably used NADH as a cofactor and had an apparent K(m) value of 0.03 mM against furfural. In the presence of 1 g liter(−1) furfural and 10% xylose (wt/vol), yghA overexpression in an ethanologenic E. coli strain SSK42 resulted in an ethanol efficiency of ∼97%, with a 5.3-fold increase in ethanol titers compared to the control. YghA also exhibited activity against the less toxic inhibitor 5-hydroxymethyl furfural, which is formed due to dehydration of hexose sugars, and thus is a formidable target for overexpression in ethanologenic strain for fermentation of sugars in biomass hydrolysate. IMPORTANCE Lignocellulosic biomass represents an inexhaustible source of carbon for second-generation biofuels. Thermo-acidic pretreatment of biomass is performed to loosen the lignocellulosic fibers and make the carbon bioavailable for microbial metabolism. The pretreatment process also results in the formation of inhibitors that inhibit microbial metabolism and increase production costs. Furfural is a potent furan inhibitor that increases the toxicity of other inhibitors present in the hydrolysate. Thus, it is desirable to engineer furfural tolerance in E. coli for efficient fermentation of hydrolysate sugars.

Climate change is known to simultaneously increase berry sugars but decrease anthocyanins, leading to an imbalance between sugars and anthocyanins in grape berries. To restore the balance of sugars and anthocyanins, carbon limitation by leaf removal and exogenous abscisic acid (ABA) were separately or simultaneously applied to Vitis vinifera cv. Cabernet Sauvignon fruit cuttings to decipher their effects on berry quality with metabolite and whole-genome transcriptome analyses. Carbon limitation decreased the hexose concentration and fully blocked the accumulation of anthocyanins. However, exogenous ABA increased the anthocyanin concentration under both carbon limitation and sufficient conditions. Carbon limitation and exogenous ABA induced the profound remodeling of the whole-genome transcriptome and altered the anthocyanin concentration by regulating the transcription levels of genes involved in the anthocyanin biosynthesis pathways as well as in the genes involved in various types of hormone signaling. Moreover, two pertinent candidate genes were identified based on the co-expression network analysis between the berry metabolite and transcriptome results, including a transcriptional factor, ERF2, and a calcineurin B-like protein-interacting protein kinase gene, CIPK25. In summary, simultaneously modifying the carbon supply by leaf removal and spraying exogenous ABA could re-establish the balance between sugars and anthocyanins to improve the qualities of grape berries via whole-genome transcriptome remodeling.

Recently, our lab found that the canonical glucose/galactose regulation pathway in yeast makes the decision to metabolize galactose based on the ratio of glucose to galactose concentrations in the external medium. This led to the question of where and how the ratio-sensing is achieved. Here, we consider the possibilities of an intracellular, extracellular, or membrane bound ratio sensing mechanisms. We show that hexose transporters in the plasma membrane are mainly responsible for glucose/galactose ratio-sensing in yeast. Further, while the glucose sensors Gpr1, Snf3, and Rgt2 are not required for ratio sensing, they help modulate the ratio sensing phenotype by regulating the expression of individual transporters in different environments. Our study provides an example of an unexpected, but potentially widespread, mechanism for making essential decisions.

Cucumber (Cucumis sativus L.), an important vegetable plant species, is susceptible to low temperature stress especially during the seedling stage. Vacuolar invertase (VI) plays important roles in plant responses to abiotic stress. However, the molecular and biochemical mechanisms of VI function in cucumber, have not yet been completely understood and VI responses to low temperature stress and it functions in cold tolerance in cucumber seedlings are also in need of exploration. The present study found that hexose accumulation in the roots of cucumber seedlings under low temperature stress is closely related to the observed enhancement of invertase activity. Our genome-wide search for the vacuolar invertase (VI) genes in cucumber identified the candidate VI-encoding gene CsVI1. Expression profiling of CsVI1 showed that it was mainly expressed in the young roots of cucumber seedlings. In addition, transcriptional analysis indicated that CsVI1 expression could respond to low temperature stress. Recombinant CsVI1 proteins purified from Pichia pastoris and Nicotiana benthamiana leaves could hydrolyze sucrose into hexoses. Further, overexpression of CsVI1 in cucumber plants could increase their hexose contents and improve their low temperature tolerance. Lastly, a putative cucumber invertase inhibitor was found could form a complex with CsVI1. In summary, these results confirmed that CsVI1 functions as an acid invertase involved in hexose accumulation and responds to low temperature stress in cucumber seedlings.

Simultaneous co-fermentation of glucose and xylose is a key desired trait of engineered Saccharomyces cerevisiae for efficient and rapid production of biofuels and chemicals. However, glucose strongly inhibits xylose transport by endogenous hexose transporters of S. cerevisiae. We identified structurally distant sugar transporters (Lipomyces starkeyi LST1_205437 and Arabidopsis thaliana AtSWEET7) capable of co-transporting glucose and xylose from previously unexplored oleaginous yeasts and plants. Kinetic analysis showed that LST1_205437 had lenient glucose inhibition on xylose transport and AtSWEET7 transported glucose and xylose simultaneously with no inhibition. Modelling studies of LST1_205437 revealed that Ala335 residue at sugar binding site can accommodates both glucose and xylose. Docking studies with AtSWEET7 revealed that Trp59, Trp183, Asn145 and Asn179 residues stabilized the sugars, allowing both xylose and glucose to be co-transported. In addition, we altered sugar preference of LST1_205437 by single amino acid mutation at Asn365. Our findings provide a new mechanistic insight on glucose and xylose transport mechanism of sugar transporters and the identified sugar transporters can be employed to develop engineered yeast strains for producing cellulosic biofuels and chemicals. This article is protected by copyright. All rights reserved.

Sugar is the key factor in the formation of fruit quality. Hexose is the main form of sugar accumulated in ordinary cultivated tomatoes, while a small number of wild-type tomato varieties can also accumulate sucrose. Although several studies have focused on wild sucrose-accumulating tomatoes, the sugar accumulation mechanism in tomato is still unclear. Here, two cherry tomatoes lines that accumulated sucrose and hexose respectively were selected to analyze the assimilates unloading pathway and sugar accumulation mechanism using CF tracing, cytological observation, proteomics methods, etc. The results indicated that the later stages of fruit development were key stages for sugar accumulation, and sucrose-accumulating (S) cherry tomatoes had higher sucrose contents in the fruits, while hexose-accumulating (H) cherry tomatoes accumulated more glucose, fructose and starch. The unloading pathway of assimilates in the S cherry tomato was switched from apoplastic to symplastic during fruit development, and the opposite was true in the H type. Plasmodesmata transport may be the main means of sucrose accumulation and high activity or expression levels of acid invertase (AI) and SUT1 may be important factors in hexose accumulation in H and S cherry tomatoes, respectively. In addition to sugar metabolism, photosynthesis, fatty acid metabolism and other secondary metabolism pathways also play important roles in sugar accumulation. This study provides detailed evidence for the tomato sugar accumulation mechanism from the perspective of cell structure, physiology and molecular biology, providing a theoretical basis for the improvement of tomato quality and aiding the utilization of tomato genetic resources.

Yeast metabolizes the disaccharide melibiose by hydrolyzing it into equimolar concentrations of glucose and galactose by MEL1-encoded α-galactosidase. Galactose metabolizing genes (including MEL1) are induced by galactose and repressed by glucose, which are the products of melibiose hydrolysis. Therefore, how melibiose catabolization and utilization take place by circumventing the glucose repression is an enigma. Other than the galactose metabolizing genes MTH1, a negative regulator of glucose signal pathway has Gal4p binding sites and is induced by galactose and repressed by high glucose concentration. But, at low or no glucose MTH1 along with its paralogue STD1 represses hexose transporters, that are involved in glucose transport. This sort of tuning of glucose and galactose regulation motivated us to delineate the role of MTH1 as a regulator of MEL1 expression and melibiose utilization. The deletion mutant of MTH1 shows growth defect on melibiose and this growth defect is enhanced upon the deletion of both MTH1 and its paralogue STD1. Microscopy and flowcytometry analysis, suggest, that even though MEL1 and GAL1 promoter are under Gal4p and Gal80p regulation, upon deletion of MTH1 it hampers only MEL1 expression, but not the GAL1 gene expression. By using 2-Deoxy galactose toxicity assay, we observed phenotypic heterogeneity in cells grown on melibiose i.e. after cleaving of melibiose a fraction of cell population utilizes glucose and another fraction utilizes galactose and coexist together. Understanding GAL/MEL gene expression patterns in melibiose will have great implication to understand various other complex sugar utilizations, tunable gene expressions and complex feedback gene regulations.SignificanceSugar metabolism is an important phenomenon to understand the regulation of gene expression. Glucose is the most preferred carbon source. Yeast follows glycolytic pathways like cancer cells for metabolism of sugars and understanding this will throw more light to the metabolism of cancer cells. In this communication we observed cell-to-cell heterogeneity in yeast cells playing a key role in metabolism of a complex disaccharide melibiose, which gets cleaved into glucose and galactose by α-galactosidase. Glucose represses α-galactosidase and galactose induces it. Because of the heterogeneous population of cells one fraction consumes glucose liberated by melibiose hydrolysis, therefore it is not sufficient to repress α-galactosidase and other GAL genes. Therefore, GAL genes are expressed and help in metabolizing melibiose and galactose.

Background Cellulose is not only a common component in vascular plants, but also has great economic benefits for paper, wood, and industrial products. In addition, its biosynthesis is highly regulated by carbohydrate metabolism and allocation in plant. MdFRK2, which encodes a key fructokinase (FRK) in apple, showed especially high affinity to fructose and regulated carbohydrate metabolism. Results It was observed that overexpression of MdFRK2 in apple decreased sucrose (Suc) and fructose (Fru) with augmented FRK activity in stems, and caused the alterations of many phenotypic traits that include increased cellulose content and an increase in thickness of the phloem region. To further investigate the involved mechanisms, we generated FRK2-OE poplar lines OE#1, OE#4 and OE#9 and discovered (1) that overexpression of MdFRK2 resulted in the huge increased cellulose level by shifting the fructose 6-phosphate or glucose 6-phsophate towards UDPG formation, (2) a direct metabolic pathway for the biosynthesis of cellulose is that increased cleavage of Suc into UDP-glucose (UDPG) for cellulose synthesis via the increased sucrose synthase (SUSY) activity and transcript levels of PtrSUSY1, (3) that the increased FRK activity increases the sink strength overall so there is more carbohydrate available to fuel increased cambial activity and that resulted in more secondary phloem. These results demonstrated that MdFRK2 overexpression would significantly changes the photosynthetic carbon flux from sucrose and hexose to UDPG for increased cellulose synthesis. Conclusions The present data indicated that MdFRK2 overexpression in apple and poplar changes the photosynthetic carbon flux from sucrose and hexose to UDPG for stem cellulose synthesis. A strategy is proposed to increase cellulose production by regulating sugar metabolism as a whole.

AMPK/SNF1 is the master regulator of energy homeostasis in eukaryotic cells and has a key role in glucose de-repression. If glucose becomes depleted, Snf1 is phosphorylated and activated. Activation of Snf1 is required but is not sufficient for mediating glucose de-repression indicating a second glucose-regulated step that adjusts the Snf1 pathway. To elucidate this regulation, we further explore the spatial dynamics of Snf1 and Mig1 and how they are controlled by concentrations of hexose sugars. We utilize fluorescence recovery after photobleaching (FRAP) to study the movement of Snf1 and how it responds to external glucose concentrations. We show that the Snf1 pathway reacts both to the presence and to the absolute concentration of glucose. Furthermore, we identify a negative feedback loop regulating Snf1 activity. We also show that Mig1 localization correlates with the Snf1 phosphorylation pattern and not with the Mig1 phosphorylation pattern, suggesting that inactivation of Snf1 has a more pronounced effect on the localization of Mig1 than on the phosphorylation of Mig1. Our data offer insight into the true complexity of regulation of this central signaling pathway by one signal (glucose depletion) interpreted by the cell in different ways.

Human milk oligosaccharides (HMOs) have attracted much attention in recent years not only as a prebiotic factor, but in particular as an essential component in infant nutrition related to their impact in innate immunity. The backbone structures of complex HMOs generally contain single or repetitive lacto-N-biose (type 1) or lactosamine (type 2) units in either linear or branched chains extending from a lactose core. While all known branched structures originate from 3,6-substitution of the lactosyl core galactose, we here describe a new class of HMOs that tentatively branch at terminal galactose of 6’-galactosyllactose. Another novel feature of this class of HMOs was found in linear oligo-galactosyl chains linked to one of the N-acetylglucosamine (GlcNAc) branches. The novel structures exhibit general formulas with hexose vs. hexosamine contents of 5/2 to 8/2 and can be designated as high-galactose (HG)-HMOs. In addition, up to three fucosyl residues are linked to the octa- to dodecasaccharides, which were detected in two human milk samples from Lewis blood group defined donors. Structural analyses of methylated glycans and their alditols comprised MALDI mass spectrometry, ESI-(CID)MS and linkage analyses by GC-MS of the derived partially methylated alditol acetates. Enzymatic degradation by application of β1-3,4-specific galactosidase supported the presence of terminal galactose linked β1−6 to one of the two GlcNAc branches.

SummaryA common cellular task is to match gene expression dynamically to a range of concentrations of a regulatory molecule. Studying glucose transport in budding yeast, we determine mechanistically how such matching occurs for seven hexose transporters. By combining time-lapse microscopy with mathematical modelling, we find that levels of transporters are history-dependent and are regulated by a push-pull system comprising two types of repressors. Repression by these two types varies with glucose in opposite ways, and not only matches the expression of transporters by their affinity to a range of glucose concentrations, but also the expression of some to how glucose is changing. We argue that matching is favoured by a rate-affinity trade-off and that the regulatory system allows yeast to import glucose rapidly enough to starve competitors. Matching expression to a pattern of input is fundamental, and we believe that push-pull repression is widespread.

Summary • Stomatal opening is caused by guard cell swelling due to an accumulation of osmotica. We investigated the release of carbon from guard cell chloroplasts as a source for the production of organic osmotica. • Photosynthetically active chloroplasts were isolated from guard cell protoplasts of Vicia faba. Export of metabolites into the surrounding medium was analyzed by silicone oil filtering centrifugation and spectrophotometrically by coupled metabolite assays. Effects of external oxaloacetate and 3-phosphoglycerate on photosynthetic electron transport were examined by recording chlorophyll fluorescence. • In the light, guard cell chloroplasts exported triose phosphates, glucose, maltose and hexose phosphates. The presence of phosphate in the medium was essential for the release of phosphorylated compounds and also strongly enhanced the export of glucose and maltose. Total efflux of carbon from illuminated guard cell chloroplasts was on average 486 µatom C (mg Chl)−1 h−1, which was significant with respect to the carbon requirement for stomatal opening. • Metabolites released by illuminated guard cell chloroplasts originated predominantly from starch breakdown. Photosynthetic electron transport provided redox power for the reduction of oxaloacetate and 3-phosphoglycerate.

Wolfberry fruit has been attracting attention for centuries in Asian countries as a traditional herbal medicine and valuable nourishing tonic. Revealing the spatial distribution changes of important endogenous molecules during plant development is of great significance for investigating the physiological roles, nutritional and potential functional values of phytochemicals in wolfberry fruit. However, their spatial distribution information during fruit development has not been extensively explored due to the lack of efficient analytical techniques. In this work, matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) was performed to visualize the spatial distribution of the endogenous molecules during fruit development. From the mass spectrum imaging, the choline, betaine and citric acid were distributed evenly throughout the entire fruit at all development stages. The hexose was distributed in the endocarp and flesh tissue, while sucrose was located in the seeds. Additionally, several phenolic acids and flavonoids were accumulated in the exocarp during fruit development, which indicated that they seemingly played protective roles in wolfberry fruit growth progress against abiotic and biotic stress. From the collected data, we found that the signal intensities of citric acid were decreased, while choline, betaine, hexose and sucrose were increased with fruit development. These results indicate that MALDI-MSI may become a favorable tool for studying of the spatial distribution and effective use of endogenous molecules, which provide a simple and intuitive way for authenticity identification, classification of drug food homologous foods and further understanding the physiological roles of endogenous molecules.

Background Caldicellulosiruptor kronotskyensis has gained interest for its ability to grow on various lignocellulosic biomass. The aim of this study was to investigate the growth profiles of C. kronotskyensis in the presence of mixtures of glucose–xylose. Recently, we characterized a diauxic-like pattern for C. saccharolyticus on lignocellulosic sugar mixtures. In this study, we aimed to investigate further whether C. kronotskyensis has adapted to uptake glucose in the disaccharide form (cellobiose) rather than the monosaccharide (glucose). Results Interestingly, growth of C. kronotskyensis on glucose and xylose mixtures did not display diauxic-like growth patterns. Closer investigation revealed that, in contrast to C. saccharolyticus, C. kronotskyensis does not possess a second uptake system for glucose. Both C. saccharolyticus and C. kronotskyensis share the characteristics of preferring xylose over glucose. Growth on xylose was twice as fast (μmax = 0.57 h−1) as on glucose (μmax = 0.28 h−1). A study of the sugar uptake was made with different glucose–xylose ratios to find a kinetic relationship between the two sugars for transport into the cell. High concentrations of glucose inhibited xylose uptake and vice versa. The inhibition constants were estimated to be KI,glu = 0.01 cmol L−1 and KI,xyl = 0.001 cmol L−1, hence glucose uptake was more severely inhibited by xylose uptake. Bioinformatics analysis could not exclude that C. kronotskyensis possesses more than one transporter for glucose. As a next step it was investigated whether glucose uptake by C. kronotskyensis improved in the form of cellobiose. Indeed, cellobiose is taken up faster than glucose; nevertheless, the growth rate on each sugar remained similar. Conclusions C. kronotskyensis possesses a xylose transporter that might take up glucose at an inferior rate even in the absence of xylose. Alternatively, glucose can be taken up in the form of cellobiose, but growth performance is still inferior to growth on xylose. Therefore, we propose that the catabolism of C. kronotskyensis has adapted more strongly to pentose rather than hexose, thereby having obtained a specific survival edge in thermophilic lignocellulosic degradation communities.

Background Candida albicans is an opportunistic commensal microorganism, often associated with severe infections in immunosuppressed individuals. C. albicans has hexose transporters that may favor the intracellular accumulation of photosensitizer (PS). the aims of this study were to investigate the influence of glucose load on photodynamic antimicrobial chemotherapy (PACT); and the role that membrane transport system plays on this therapy in the presence of glucose. Material and methods Strains of C. albicans were selected: ATCC 10231, YEM 12, YEM 13, YEM 14 and YEM 15. All strains were grown aerobically on Sabouraud agar and incubated at 30 °C for 24 h. The strains were treated with and without glucose, and divided into Control (no treatment), LED light (660 nm, 166 mW/cm2), Photosensitizer (100 μM methylene blue) and PACT at 1, 3 and 6 min of irradiation groups. The colony forming units were counted and data submitted to statistical analysis (ANOVA) and Tukey's test. The concentration of methylene blue (MB) outside the yeast was measured by fluorescence spectroscopy. Results PACT inactivate C. albicans and the presence of glucose did not affect the killing effect for most strains. Only YEM12 was partially affected by its presence. Regarding efflux systems, ABC overexpressing strain showed a protective effect on the yeast cells. We observed that yeast with overexpression of major facilitator superfamily (MFS) membrane pore tended to accumulate more MB in its cytoplasm, whereas strains that overexpressed ABC pumps (ATP-binding cassette transporters) tended to decrease MB uptake and survive the photodynamic challenge. Conclusion Presence of glucose showed a small effect on PACT . The accumulation of MB on yeast induces more photodynamic inactivation; however, the photodynamic efficacy depends on the type and characteristics of the microbial strain.

The objective of this study was to evaluate the effects of hydraulic retention time (HRT) (8–1 h) on H 2 production from sugarcane juice (5000 mg COD L −1 ) in mesophilic (30 °C, AFBR-30) and thermophilic (55 °C, AFBR-55) anaerobic fluidized bed reactors (AFBRs). At HRTs of 8 and 1 h in AFBR-30, the H 2 production rates were 60 and 116 mL H 2 h −1 L −1 , the hydrogen yields were 0.60 and 0.10 mol H 2 mol −1 hexose, and the highest bacterial diversities were 2.47 and 2.34, respectively. In AFBR-55, the decrease in the HRT from 8 to 1 h increased the hydrogen production rate to 501 mL H 2 h −1 L −1 at the HRT of 1 h. The maximum hydrogen yield of 1.52 mol H 2 mol −1 hexose was observed at the HRT of 2 h and was associated with the lowest bacterial diversity (0.92) and highest bacterial dominance (0.52).

Although hexoses glucose and fructose serve as important energy sources of food, up to now, there is little information about hexose transporters in birds’ intestinal epithelium during their first postnatal week. The aim of the investigation was to carry out an immunohistochemical study of integral membrane proteins glucose transporter-2 and -5 (GLUT-2 and GLUT-5) on intestinal epithelial cells of ostrich chicks during their first postnatal week. The material from duodenum and ileum was collected from 9 female ostriches (Struthio camelus var. Domesticus) divided into three age groups, three birds in each group: chicks immediately after hatching, 3-day-old ostriches and 7-day-old chicks. The material was fixed in 10% formalin, embedded into paraffin, slices 7μm thick were cut followed by immunohistochemical staining with polyclonal primary antibodies Rabbit anti- GLUT-2 and Rabbit anti-GLUT-5, carried out according to the manufacturer’s guidelines (IHC kit, Abcam, UK). Immunohistochemical localization of GLUT-2 and -5 in the intestinal epithelial cells in ostriches of different age groups was determined. In the groups of chicks after hatching and 3-day-old ostriches, enterocytes in duodenal epithelium were mostly unstained and goblet cells stained weakly for both antibodies. Weak staining of enterocytes and goblet cells was also noted in the ileal epithelium of the chick after hatching. Moderate staining of goblet cells was noted in the 3-day-old chicks’ ileal epithelium. In 7-day-old ostriches, the expression of both antibodies was weak in duodenal but moderate in ileal epithelial cells. The pattern of immunohistochemical expression of GLUT-2 and GLUT-5 in ostriches’ intestinal epithelial cells confirms our hypothesis that the intestinal tract of ostriches after hatching is not yet entirely capable of transportation of hexoses and showed that it is completing gradually during the first postnatal week.

Hexokinase (HXK) catalyses hexose phosphorylation and senses glucose signalling to control carbon partition in plants. In sugarcane, HXK is involved in regulating source/sink carbon balance. However, the characteristics of sugarcane HXK genes remain unclear. This study revealed that the full-length cDNA sequence of the HXK gene ShHXK8 cloned from sugarcane mature leaves contains a 1560 bp open reading frame and encodes a putative protein of 519 amino acids. Amongst HXKs from monocot and dicot subjected to phylogenetic analysis, ShHXK8 was clustered into a monocot-specific group with OsHXK7, SbHXK8 and ZmHXK1. Gene expression analysis in Saccharum spp. hybrid ROC22 plants showed that ShHXK8 was expressed in leaves and stalks and expressed highly in mature leaves and young internodes. The difference in the expression of ShHXK8 was compared in high-sugar-content (HSC) and low-sugar-content (LSC) Saccharum officinarum plants. The expression of ShHXK8 in the leaves and internodes 3–24 of HSC plants was lower than that of LSC plants. The expression of ShHXK8 in the internodes 1–2 of HSC plants was higher than that of LSC plants. ShHXK8–GFP fusion was located in the cytosol by transiently expressing the corresponding vectors in rice mesophyll protoplasts. ShHXK8 might function as a cytoplasmic hexokinase involved in hexose metabolism in sugarcane mature leaves and young internodes.

Carrageenase is useful for preparation of carrageenan oligosaccharides, which have significant bioactivity. We expressed a κ‑carrageenase gene from Zobellia sp. ZL-4 in full-length (κ-ZL-4) or after truncation of the carbohydrate binding module and the Type-IX secretion module (κ-ZL-4-GH16). κ-ZL-4-GH16 showed a specific activity (134.22 U/mg) 1.93 times higher than that of κ-ZL-4, and its thermal and pH stability also increased. The best activity of κ-ZL-4-GH16 was presented at pH 3.0–6.0, which was lower than the optimal pH of reported κ-carrageenases. The enzyme-substrate affinity of κ-ZL-4-GH16 was higher than that of κ-ZL-4, demonstrated by its lower Michaelis-Menten constant (0.704 mg/mL at pH 6.0). Importantly, κ-ZL-4-GH16 released 10-fold more κ-carrageenan disaccharides than κ-ZL-4. The κ-carrageenan tetrose and hexose produced by the two enzymes were purified and structurally identified. Molecular docking with κ-carrageenan hexose suggested that the efficiency improvement after truncation might be attributed to the conformation differences of the two enzymes.

Over-express galactokinase (Galk1) in tissue plasminogen activator (tPA) producing CHO cells as a potential strategy to improve cell growth and product synthesis. tPA producing CHO cells were transfected with the galactokinase (Galk1) gene. CHO-Galk1 cells showed a 39% increase of the specific growth rate in galactose. Moreover, clones were able to use this hexose as their main carbon source to sustain growth contrary to their parental cell line. Metabolic Flux Analysis revealed that the CHO-Galk1 selected clone shows an active metabolism towards biomass and product synthesis, characterized by higher fluxes in the TCA cycle, which is consistent with increased cellular densities and final product concentration. This cellular engineering strategy, where modifications of key points of alternative carbon sources metabolism lead to an improved metabolism of these sugars, is a starting point towards the generation of new cell lines with reduced lactate synthesis and increased cell growth and productivity.

Silencing expression of SlSWEET1a substantially reduced hexose accumulation in young leaves of tomato plants, revealing the key role of SlSWEET1a in glucose import to sink leaves., Sugar allocation from source to sink (young) leaves, critical for plant development, relies on activities of plasma membrane sugar transporters. However, the key sugar unloading mechanism to sink leaves remains elusive. SWEET transporters mediate sugar efflux into reproductive sinks; therefore, they are promising candidates for sugar unloading during leaf growth. Transcripts of SlSWEET1a, belonging to clade I of the SWEET family, were markedly more abundant than those of all other 30 SlSWEET genes in young leaves of tomatoes. High expression of SlSWEET1a was also detected in reproductive sinks, such as flowers. SlSWEET1a was dominantly expressed in leaf unloading veins, and the green fluorescent protein (GFP) fusion protein was localized to the plasma membrane using Arabidopsis protoplasts, further implicating this carrier in sugar unloading. In addition, yeast growth assays and radiotracer uptake analyses further demonstrated that SlSWEET1a acted as a low-affinity (Km ~100 mM) glucose-specific carrier with a passive diffusion manner. Finally, virus-induced gene silencing of SlSWEET1a expression reduced hexose accumulation to ~50% in young leaves, with a parallel 2-fold increase in mature leaves. Thus, we propose a novel function for SlSWEET1a in the uptake of glucose into unloading cells as part of the sugar unloading mechanism in sink leaves of tomato.