Effects of priming glycosyltransferase genes cps 2E and cps 4E on the exopolysaccharide biosynthesis by Lactiplantibacillus plantarum YM-4-3 strain.

Microbial exopolysaccharides (EPS) have various physiological functions such as antioxidant, anti-tumor, cholesterol lowering, and immune regulation. However, improving traditional fermentation conditions to increase the production of EPS from Lactiplantibacillus plantarum (L. plantarum) is limited. In this study, we aimed to better improve EPS production and physiological functions of L. plantarum YM-4-3 strain by overexpressing and knocking out the priming glycosyltransferase genes cps 2E and cps 4E for the first time. As a result, the EPS production of the overexpression strain was 30.15 %, 26.84 % and 36.29 % higher than WT, respectively. The EPS production of the knockout strain was significantly lower than that of the WT. At the same time, transcriptome data showed that the gene expression levels of each experimental strain had changed. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways found that the glycolysis/gluconeogenesis pathway had the highest gene enrichment in the metabolic pathway. The monosaccharide components of the EPS of each experimental strain were different from those of the WT and the EPS of the experimental strain showed stronger activity against oxidation. In conclusion, this study contributes to the efficient production and application of L. plantarum EPS and helps to understand the mechanism of EPS regulation in L. plantarum.

Overexpression of tomato glutathione reductase (SlGR) in transgenic tobacco enhances salt tolerance involving the S-nitrosylation of GR.

S-nitrosylation, a post-translational modification (PTM) dependent on nitric oxide, is essential for plant development and environmental responsiveness. However, the function of S-nitrosylation of glutathione reductase (GR) in tomato (SlGR) under NaCl stress is yet uncertain. In this study, sodium nitroprusside (SNP), an exogenous NO donor, alleviated the growth inhibition of tomato under NaCl treatment, particularly at 100 µM. Following NaCl treatment, the transcripts, enzyme activity, and S-nitrosylated level of GR were increased. In vitro, the SlGR protein was able to be S-nitrosylated by S-nitrosoglutathione (GSNO), significantly increasing the activity of GR. SlGR overexpression transgenic tobacco plants exhibited enhanced germination rate, fresh weight, and increased root length in comparison to wild-type (WT) seedlings. The accumulation of reactive oxygen species (ROS) was lower, whereas the expression and activities of GR, ascorbate peroxidase (APX), superoxide dismutase (SOD), and catalase (CAT); the ratio of ascorbic acid/dehydroascorbic acid (AsA/DHA), reduced glutathione/oxidized glutathione (GSH/GSSG), total soluble sugar and proline contents; and the expression of stress-related genes were higher in SlGR overexpression transgenic plants in comparison to the WT plants following NaCl treatment. The accumulation of NO and S-nitrosylated levels of GR in transgenic plants was higher in comparison to WT plants following NaCl treatment. These results indicated that S-nitrosylation of GR played a significant role in salt tolerance by regulating the oxidative state.

Highly Efficient Biotransformation of Phenolic Glycosides Using a Recombinant ß-Glucosidase From White Rot Fungus Trametes trogii.

Phenolic glycosides are the important bioactive molecules, and their bioavailability can be influenced by enzyme hydrolysis, such as ß-glucosidases (EC3.2.1.21) and other glycosyl hydrolases (GHs). Wood rotting fungi possess a superfamily of GHs, but little attention has been paid to the GHs and their potential applications in biotransformation of phenolic glycosides. In this study, two GH3 gene family members of Trametes trogii S0301, mainly expressed in the carbon sources conversion stage were cloned, and TtBgl3 coded by T_trogii_12914 showed ß-glucosidase activity toward 4-nitrophenyl ß-D-glucopyranoside (pNPG). The recombinant TtBgl3 preferred an intermediately neutral optimum pH with >80% of the maximum activity at pH 5.0-7.0 and was stable at a wide range of pH (5.0-10.0). Phenolic glycosides transformation experiments showed that TtBgl3 was a dual-activity enzyme with both activities of aryl-ß-D-glucosidase and ß-glucuronidase, and could hydrolyze the ß-glucoside/glucuronide bond of phenolic glycosides. Under optimized conditions, the recombinant TtBgl3 had much higher transformation efficiency toward the ß-glucoside bond of gastrodin, esculin and daidzin than ß-glucuronide bond of baicalin, with the transformation rate of 100 and 50%, respectively. Our homology modeling, molecular docking, and mutational analysis demonstrated that His85 and Lys467 in the acceptor-binding pocket of TtBgl3 were the potential active sites. The point mutation of His85 and Lys467 leads to the significantly impaired catalytic activity toward pNPG and also the weak transformation efficiency toward gastrodin. These findings provide insights for the identification of novel GH3 ß-glucosidases from T. trogii and other wood-rotting fungi. Furthermore, TtBgl3 might be applied as green and efficient biological catalysts in the deglycosylation of diverse phenolics to produce bioactive glycosides for drug discovery in the future.

A Putative Guanosine Triphosphate Cyclohydrolase I Named CaGCH1 Is Involved in Hyphal Branching and Fruiting Development in Cyclocybe aegerita.

Guanosine triphosphate (GTP) cyclohydrolase I (GCH1) is the limiting enzyme of the tetrahydrobiopterin (BH4) synthesis pathway. The disruption of gch1 gene may cause conditional lethality due to folic acid auxotrophy in microorganisms, although the function of gch1 in basidiomycetes has not been deciphered so far. In the present study, gch1 expression in Cyclocybe aegerita (cagch1) was downregulated using the RNAi method, which resulted in growth retardation in both solid and liquid medium, with the hyphal tips exhibiting increased branching compared to that in the wild strain. The development of fruiting bodies in the mutant strains was significantly blocked, and there were short and bottle-shaped stipes. The transcriptional profile revealed that the genes of the MAPK pathway may be involved in the regulation of these effects caused by cagch1 knockdown, which provided an opportunity to study the role of gch1 in the development process of basidiomycetes.

Overexpression of tomato thioredoxin h (SlTrxh) enhances excess nitrate stress tolerance in transgenic tobacco interacting with SlPrx protein.

The thioredoxin (Trx) system plays a vital function in cellular antioxidative defense. However, little is known about Trx in tomato under excess nitrate. In this study, we isolated the tomato gene encoding h-type Trx gene (SlTrxh). The mRNA transcript of SlTrxh in roots and leaves of tomato was induced incrementally under excess nitrate for 24 h. Subcellular localization showed that SlTrxh might localize in the cytoplasm, nucleus and plasma membrane. Enzymatic activity characterization revealed that SlTrxh protein possesses the disulfide reductase function and Cysteine (Cys) 54 is important for its activity. Overexpressing SlTrxh in tobacco resulted in increasing seed germination rate, root length and decreasing H2O2 and O2- accumulation, compared with the wild type (WT) tobacco under nitrate stress. While overexpressing SlTrxhC54S (Cysteine 54 mutated to Serine) in tobacco showed decreased germination rate and root length compared with the WT after nitrate treatment. After nitrate stress treatment, SlTrxh overexpressing transgenic tobacco plants have lower malonaldehyde (MDA), H2O2 contents and Reactive Oxygen Species (ROS) accumulation, and higher mRNA transcript level of NtP5CS, NtDREB2, higher ratio of ASA/DHA and GSH/GSSG, higher activities of ascorbate peroxidase and NADP thioredoxin reductase. Besides, SlTrxh overexpressing plants showed higher tolerance to Methyl Viologen (MV) in the seed germination and seedling stage. The yeast two-hybrid, pull-down, Co-immunoprecipitation and Bimolecular luciferase complementation assay confirmed that SlTrxh physically interacted with tomato peroxiredoxin (SlPrx). These results suggest that SlTrxh contributes to maintaining ROS homeostasis under excess nitrate stress interacting with SlPrx and Cys54 is important for its enzyme activity.

The Manganese Peroxidase Gene Family of Trametes trogii: Gene Identification and Expression Patterns Using Various Metal Ions under Different Culture Conditions.

Manganese peroxidases (MnPs), gene family members of white-rot fungi, are necessary extracellular enzymes that degrade lignocellulose and xenobiotic aromatic pollutants. However, very little is known about the diversity and expression patterns of the MnP gene family in white-rot fungi, especially in contrast to laccases. Here, the gene and protein sequences of eight unique MnP genes of T. trogii S0301 were characterized. Based on the characteristics of gene sequence, all TtMnPs here belong to short-type hybrid MnP (type I) with an average protein length of 363 amino acids, 5-6 introns, and the presence of conserved cysteine residues. Furthermore, analysis of MnP activity showed that metal ions (Mn2+ and Cu2+) and static liquid culture significantly influenced MnP activity. A maximum MnP activity (>14.0 U/mL) toward 2,6-DMP was observed in static liquid culture after the addition of Mn2+ (1 mM) or Cu2+ (0.2 or 2 mM). Moreover, qPCR analysis showed that Mn2+ obviously upregulated the Group I MnP subfamily (T_trogii_09901, 09904, 09903, and 09906), while Cu2+ and H2O2, along with changing temperatures, mainly induced the Group II MnP subfamily (T_trogii_11984, 11971, 11985, and 11983), suggesting diverse functions of fungal MnPs in growth and development, stress response, etc. Our studies here systematically analyzed the gene structure, expression, and regulation of the TtMnP gene family in T. trogii, one of the important lignocellulose-degrading fungi, and these results extended our understanding of the diversity of the MnP gene family and helped to improve MnP production and appilications of Trametes strains and other white-rot fungi.

Alternative Splicing of Heat Shock Transcription Factor 2 Regulates the Expression of Laccase Gene Family in Response to Copper in Trametes trogii.

White-rot fungi, especially Trametes strains, are the primary source of industrial laccases in bioenergy and bioremediation. Trametes strains express members of the laccase gene family with different physicochemical properties and expression patterns. However, the literature on the expression pattern of the laccase gene family in T. trogii S0301 and the response mechanism to Cu2+, a key laccase inducer, in white-rot fungal strains is scarce. In the present study, we found that Cu2+ could induce the mRNAs and proteins of the two alternative splicing variants of heat shock transcription factor 2 (TtHSF2). Furthermore, the overexpression of alternative splicing variants TtHSF2α and TtHSF2ß-I in the homokaryotic T. trogii S0301 strain showed opposite effects on the extracellular total laccase activity, with the maximum laccase activity of approximately 0.6 U mL-1 and 3.0 U mL-1, respectively, on the eighth day, which is 0.4 and 2.3 times that of the wild type strain. Similarly, TtHSF2α and TtHSF2ß-I play opposite roles in the oxidation tolerance to H2O2 In addition, the direct binding of TtHSF2α to the promoter regions of the representative laccase isoenzymes (TtLac1 and TtLac13) and protein-protein interactions between TtHSF2α and TtHSF2ß-I were detected. Our results demonstrate the crucial roles of TtHSF2 and its alternative splicing variants in response to Cu2+ We believe that these findings will deepen our understanding of alternative splicing of HSFs and their regulatory mechanism of the laccase gene family in white-rot fungi.Importance The members of laccase gene family in Trametes strains are the primary source of industrial laccase and have gained widespread attention. Increasing the yield and enzymatic properties of laccase through various methods has always been a topic worthy of attention, and there is no report on the regulation of laccase expression through HSF transcription factor engineering. Here, we found that two alternative splicing variants of TtHSF2 functioned oppositely in regulating the expression of laccase genes, and copper can induce the expression of almost all members of the laccase gene family. Most importantly, our study suggested that TtHSF2 and its alternative splicing variants are vital for copper-induced production of laccases in T. trogii S0301.

Transcriptome analysis of Cryptococcus humicola under aluminum stress revealed the potential role of the cell wall in aluminum tolerance.

Aluminum (Al) toxicity is one of the most important limiting factors for crop yield in acidic soils. Bound Al gets converted into a toxic ionic state (Al3+) in acidic soil. Recent studies have shown that Al can act on the cell walls, cell membranes, organelles, and nuclei of microorganisms and affect substance and energy metabolism. To explore the gene expression at the transcriptional level under Al stress, we sequenced the transcriptome of Cryptococcus humicola, which is a highly Al-resistant yeast strain isolated from acidic soil and tolerates up to 200 mM Al3+. The screening conditions for genes from the control and experimental group were a false discovery rate (FDR) <0.05 and log 2|FC| > 1. A total of 4760 genes were differentially expressed, among which 3066 were upregulated and 1694 were downregulated. These genes control glycometabolism, protein synthesis, lipid metabolism and signalling pathways. Eleven selected differentially expressed genes were further validated using qRT-PCR. The results suggested that Al stress leads to complex responses in C. humicola. The effects of Al on the ß-d-glucan and mannose contents and Al accumulation in the cell wall were determined. With an increase in the Al treatment time and concentration, the contents of ß-d-glucan and mannose showed a trend of first increasing and then decreasing. Under Al treatment, the Al content of the cell wall also showed a trend of first increasing and then decreasing. These results suggested that Al accumulates in the cell wall and the cell wall plays a vital role in the Al resistance of C. humicola. The differentially expressed genes provide a foundation for the further study of Al tolerance in C. humicola.

A Thermo-Active Laccase Isoenzyme From Trametes trogii and Its Potential for Dye Decolorization at High Temperature.

A thermo-activation and thermostable laccase isoenzyme (Lac 37 II) produced by Trametes trogii S0301 at 37°C was purified to apparent homogeneity by anionic exchange chromatography and sephadex G-75 chromatography, with 12.3% of yeiled and a specific activity of 343.1 U mg-1. The molecular weight of the purified Lac 37 II was estimated to be approximately 56 kDa in 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The optimal pH and temperature for the protein was 2.7 and 60°C, respectively. The purified Lac 37 II showed higher resistance to all tested metal ions and organic solvents except for Fe2+ and Cd2+ at 37°C and the activity of the purified Lac 37 was significantly enhanced by Cu2+ at 50 mM. The K cat , K m , and K cat /K m of Lac 37 II were 2.977 s-1, 16.1 µM, and 184.9 s-1 µM-1, respecively, in the condition of pH 2.7 and 60°C using ABTS as a substrate. Peptide-mass fingerprinting analysis showed that the Lac 37 II matched to the gene-deduced sequences of lcc3 in T. trogii BAFC 463, other than Lcc1, Lcc 2, and Lcc 4. Compared with laccase prepared at 28°C, the onset of thermo-activation of Lac 37 II activity occurred at 30°C with an increase of 10%, and reached its maximum at the temperatures range of 40-60°C with an increase of about 40% of their original activity. Furthermore, Lac 37 II showed the efficient decolorization ability toward triphenylmethane dyes at 60°C, with decolorization rates of 100 and 99.1% for 25 mg L-1 malachite and crystal violet in 5 h, respectively, when hydroxybenzotriazole (HBT) was used as a mediator. In conclusion, it is the first time to report a thermo-activation laccase from a thermophilic T. trogii strain, which has a better enzyme property and higher decolorization ability among fungal laccases, and it also has a further application prospective in the field of biotechnology.

Lignin degradation potential and draft genome sequence of Trametes trogii S0301.

BACKGROUND: Trametes trogii is a member of the white-rot fungi family, which has a unique ability to break down recalcitrant lignin polymers to CO2 and water, and they have enormous potential to biodegrade a wide range of toxic environmental pollutants. Because of its industrial potential, the identification of lignin-degrading enzyme systems in Trametes is an important area of research. Development and utilization of industrial value genes are suffering due to deficiency knowledge of genome available for their manipulation. RESULTS: In the present study, Homokaryotic strains of T. trogii S0301 were screened and sequencing by PacBio Sequel II platform. The final draft genome is ~ 39.88 Mb, with a contig N50 size of 2.4 Mb, this was the first genome sequencing and assembly of T. trogii species. Further analyses predicted 14,508 protein-coding genes. Results showed that T. trogii S0301 contains 602 genes encoding CAZymes, include 211 glycoside hydrolase and 117 lignin-degrading family genes, nine laccases related genes. Small subunit ribosomal RNA gene (18S rRNA) sequencing confirms its phylogenetic position. Moreover, T. trogii S0301 has the largest number of cytochromes P450 (CYPs) superfamily genes compare to other fungi. All these results are consistent with enzymatic assays and transcriptome analysis results. We also analyzed other genome characteristics in the T. trogii S0301genome. CONCLUSION: Here, we present a nearly complete genome for T. trogii S0301, which will help elucidate the biosynthetic pathways of the lignin-degrading enzyme, advancing the discovery, characterization, and modification of novel enzymes from this genus. This genome sequence will provide a valuable reference for the investigation of lignin degradation in the Trametes genus.

Endophytic fungal community of Dysphania ambrosioides from two heavy metal-contaminated sites: evaluated by culture-dependent and culture-independent approaches.

Endophytic fungal communities of Dysphania ambrosioides, a hyperaccumulator growing at two Pb-Zn-contaminated sites, were investigated through culture-dependent and culture-independent approaches. A total of 237 culturable endophytic fungi (EF) were isolated from 368 tissue (shoot and roots) segments, and the colonization rate (CR) ranged from 9.64% to 65.98%. The isolates were identified to 43 taxa based on morphological characteristics and rDNA ITS sequence analysis. Among them, 13 taxa (30.23%) were common in plant tissues from both sites; however, dominant EF were dissimilar. In culture-dependent study, 1989 OTUs were obtained through Illumina Miseq sequencing, and dominant EF were almost same in plant tissues from both sites. However, some culturable EF were not observed in total endophytic communities. We suggest that combination of both culture-dependent and culture-independent methods will provide more chances for the precise estimation of endophytic fungal community than using either of them. The tissue had more influence on the culturable fungal community structure, whereas the location had more influence on the total fungal community structure (including culturable and unculturable). Both culture-dependent and culture-independent studies illustrated that endophytic fungal communities of D. ambrosioides varied across the sites, which suggested that HM concentration of the soil may have some influence on endophytic fungal diversity.

Purification and characterization of alginate lyase from Sphingomonas sp. ZH0.

Alginate lyases degrade alginate in a beta-elimination reaction to produce oligosaccharides. Thus, alginate lyases are widely used in the food/pharmaceutical industries and are commercially valuable. In this study, four alginate lyase encoding genes were successfully cloned from Sphingomonas sp. ZH0. The expression systems of these alginate lyases were then constructed in Escherichia coli cells. The recombinant ZH0-I, ZH0-II, ZH0-III and ZH0-IV were purified from E. coli cells and were confirmed to be monomeric enzymes with molecular weights of approximately 91, 52, 67, and 113 kDa, respectively. The conditions for enzymes to have the highest specific lyase activities were 53.2 U/mg, 42 °C, pH 7.0 for ZH0-I, 103.9 U/mg, 47 °C, pH 6.5 for ZH0-II, 13.7 U/mg, 52 °C, pH 7.5 for ZH0-III, and 12.3 U/mg, 37 °C, pH 7.0 for ZH0-IV, respectively. These recombinant enzymes were stable over a pH range. Moreover, the enzymes were active in the absence of salt ions, and their activities were substantially reduced by the addition of HgCl2. ZH0-I, ZH0-II and ZH0-III belong to endotype alginate lyases, while ZH0-IV is an exotype alginate lyase. All types could degrade both poly-ß-d-mannuronate and poly-α-l-guluronate blocks, yielding alginate oligosaccharides as the main product. The Km and Vmax values were 0.51 mg/ml and 56.18 U/ml for ZH0-I, 0.47 mg/ml and 27.5 U/ml for ZH0-II, 0.55 mg/ml and 60.24 U/ml for ZH0-III, and 0.41 mg/ml and 5.53 U/ml for ZH0-IV, respectively. These features indicate that these alginate lyases are promising candidates for producing antioxidants from alginates in industrial applications.

Physiological response and transcription profiling analysis reveals the role of H2S in alleviating excess nitrate stress tolerance in tomato roots.

Soil secondary salinization caused by excess nitrate addition is one of the major obstacles in greenhouse vegetable production. Excess nitrate inhibited the growth of tomato plants, while application of 100 µM H2S donor NaHS efficiently increased the plant height, fresh and dry weight of shoot and root, root length, endogenous H2S contents and L-cysteine desulfhydrases activities. NaHS altered the oxidative status of nitrate-stressed plants as inferred by changes in reactive oxygen species (ROS) accumulation and lipid peroxidation accompanied by regulation of the activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX). Besides, NaHS increased the nitric oxide (NO) and total S-nitrosothiols (SNOs) contents, nitrate reductase (NR) activities and decreased the S-nitrosoglutathione reductase (GSNOR) activities under nitrate stress. Furthermore, microarray analysis using the Affymetrix Tomato GeneChip showed that 5349 transcripts were up-regulated and 5536 transcripts were down-regulated under NaHS and excess nitrate stress treatment, compared to the excess nitrate stress alone. The differentially expressed genes (log2 fold change >2 or <  -2) of up-regulated (213) and down-regulated (271) genes identified were functionally annotated and subsequently classified into 9 functional categories. These categories included metabolism, signal transduction, defence response, transcription factor, protein synthesis and protein fate, transporter, cell wall related, hormone response, cell death, energy and unknown proteins. Our study suggested exogenous NaHS might enhance excess nitrate stress tolerance of tomato plants by modulating ROS and reactive nitrogen species (RNS) signaling and downstream transcriptional adjustment, such as defence response, signal transduction and transcription factors.

Influence of culture media, pH and temperature on growth and bacteriocin production of bacteriocinogenic lactic acid bacteria.

There has been continued interest in bacteriocins research from an applied perspective as bacteriocins have potential to be used as natural preservative. Four bacteriocinogenic lactic acid bacteria (LAB) strains of Lactobacillus curvatus (Arla-10), Enterococcus faecium (JFR-1), Lactobacillus paracasei subsp. paracasei (JFR-5) and Streptococcus thermophilus (TSB-8) were previously isolated and identified in our lab. The objective of this study was to determine the optimal growth conditions for both LAB growth and bacteriocins production. In this study, various growth conditions including culture media (MRS and BHI), initial pH of culture media (4.5, 5.5, 6.2, 7.4 and 8.5), and incubation temperatures (20, 37 and 44 °C) were investigated for LAB growth measured as optical density (OD), bacteriocin activity determined as arbitrary unit and viability of LAB expressed as log CFU ml-1. Growth curves of the bacteriocinogenic LAB were generated using a Bioscreen C. Our results indicated that Arla-10, JFR-1, and JFR-5 strains grew well on both MRS and BHI media at growth temperature tested whereas TSB-8 strain, unable to grow at 20 °C. LAB growth was significantly affected by the initial pH of culture media (p < 0.001) and the optimal pH was found ranging from 6.2 to 8.5. Bacteriocin activity was significantly different in MRS versus BHI (p < 0.001), and the optimal condition for LAB to produce bacteriocins was determined in MRS broth, pH 6.2 at 37 °C. This study provides useful information on potential application of bacteriocinogenic LAB in food fermentation processes.

Overexpression of spinach non-symbiotic hemoglobin in Arabidopsis resulted in decreased NO content and lowered nitrate and other abiotic stresses tolerance.

A class 1 non-symbiotic hemoglobin family gene, SoHb, was isolated from spinach. qRT-PCR showed that SoHb was induced by excess nitrate, polyethylene glycol, NaCl, H2O2, and salicylic acid. Besides, SoHb was strongly induced by application of nitric oxide (NO) donor, while was suppressed by NO scavenger, nitrate reductase inhibitor, and nitric oxide synthase inhibitor. Overexpression of SoHb in Arabidopsis resulted in decreased NO level and sensitivity to nitrate stress, as shown by reduced root length, fresh weight, the maximum photosystem II quantum ratio of variable to maximum fluorescence (Fv/Fm), and higher malondialdehyde contents. The activities and gene transcription of superoxide dioxidase, and catalase decreased under nitrate stress. Expression levels of RD22, RD29A, DREB2A, and P5CS1 decreased after nitrate treatment in SoHb-overexpressing plants, while increased in the WT plants. Moreover, SoHb-overexpressing plants showed decreased tolerance to NaCl and osmotic stress. In addition, the SoHb-overexpression lines showed earlier flower by regulating the expression of SOC, GI and FLC genes. Our results indicated that the decreasing NO content in Arabidopsis by overexpressing SoHb might be responsible for lowered tolerance to nitrate and other abiotic stresses.

Laccase produced by a thermotolerant strain of Trametes trogii LK13.

Thermophilic and thermotolerant micro-organisms strains have served as the natural source of industrially relevant and thermostable enzymes. Although some strains of the Trametes genus are thermotolerant, few Trametes strains were studied at the temperature above 30 °C until now. In this paper, the laccase activity and the mycelial growth rate for Trametes trogii LK13 are superior at 37 °C. Thermostability and organic cosolvent tolerance assays of the laccase produced at 37 °C indicated that the enzyme possessed fair thermostability with 50% of its initial activity at 80 °C for 5 min, and could remain 50% enzyme activity treated with organic cosolvent at the concentration range of 25%-50% (v/v). Furthermore, the test on production of laccase and lignocellulolytic enzymes showed the crude enzymes possessed high laccase level (1000 U g (-1) ) along with low cellulose (2 U g (-1) ) and xylanase (140 U g (-1) ) activity. Thus, T. trogii LK13 is a potential strain to be applied in many biotechnological processes.

Bioethanol production from sodium hydroxide/hydrogen peroxide-pretreated water hyacinth via simultaneous saccharification and fermentation with a newly isolated thermotolerant Kluyveromyces marxianu strain.

In this study, bioethanol production from NaOH/H2O2-pretreated water hyacinth was investigated. Pretreatment of water hyacinth with 1.5% (v/v) H2O2 and 3% (w/v) NaOH at 25 °C increased the production of reducing sugars (223.53 mg/g dry) and decreased the cellulose crystallinity (12.18%), compared with 48.67 mg/g dry and 22.80% in the untreated sample, respectively. The newly isolated Kluyveromyces marxianu K213 showed greater ethanol production from glucose (0.43 g/g glucose) at 45 °C than did the control Saccharomyces cerevisiae angel yeast. The maximum ethanol concentration (7.34 g/L) achieved with K. marxianu K213 by simultaneous saccharification and fermentation (SSF) from pretreated water hyacinth at 42 °C was 1.78-fold greater than that produced by angel yeast S. cerevisiae at 30 °C. The present work demonstrates that bioethanol production achieved via SSF of NaOH/H2O2-pretreated water hyacinth with K. marxianu K213 is a promising strategy to utilize water hyacinth biomass.

Laccase produced by a thermotolerant strain of Trametes trogii LK13

Thermophilic and thermotolerant micro-organisms strains have served as the natural source of industrially relevant and thermostable enzymes. Although some strains of the Trametes genus are thermotolerant, few Trametes strains were studied at the temperature above 30 °C until now. In this paper, the laccase activity and the mycelial growth rate for Trametes trogii LK13 are superior at 37 °C. Thermostability and organic cosolvent tolerance assays of the laccase produced at 37 °C indicated that the enzyme possessed fair thermostability with 50% of its initial activity at 80 °C for 5 min, and could remain 50% enzyme activity treated with organic cosolvent at the concentration range of 25%–50% (v/v). Furthermore, the test on production of laccase and lignocellulolytic enzymes showed the crude enzymes possessed high laccase level (1000 U g−1) along with low cellulose (2 U g−1) and xylanase (140 U g−1) activity. Thus, T. trogii LK13 is a potential strain to be applied in many biotechnological processes.

UDP-glucosyltransferase71c5, a major glucosyltransferase, mediates abscisic acid homeostasis in Arabidopsis.

Abscisic acid (ABA) plays a key role in plant growth and development. The effect of ABA in plants mainly depends on its concentration, which is determined by a balance between biosynthesis and catabolism of ABA. In this study, we characterize a unique UDP-glucosyltransferase (UGT), UGT71C5, which plays an important role in ABA homeostasis by glucosylating ABA to abscisic acid -: glucose ester (GE) in Arabidopsis (Arabidopsis thaliana). Biochemical analyses show that UGT71C5 glucosylates ABA in vitro and in vivo. Mutation of UGT71C5 and down-expression of UGT71C5 in Arabidopsis cause delay in seed germination and enhanced drought tolerance. In contrast, overexpression of UGT71C5 accelerates seed germination and reduces drought tolerance. Determination of the content of ABA and ABA-GE in Arabidopsis revealed that mutation in UGT71C5 and down-expression of UGT71C5 resulted in increased level of ABA and reduced level of ABA-GE, whereas overexpression of UGT71C5 resulted in reduced level of ABA and increased level of ABA-GE. Furthermore, altered levels of ABA in plants lead to changes in transcript abundance of ABA-responsive genes, correlating with the concentration of ABA regulated by UGT71C5 in Arabidopsis. Our work shows that UGT71C5 plays a major role in ABA glucosylation for ABA homeostasis.

Voltage-dependent anion channel 2 of Arabidopsis thaliana (AtVDAC2) is involved in ABA-mediated early seedling development.

The voltage-dependent anion channel (VDAC) is the major transport protein in the outer membrane of mitochondria and plays crucial roles in energy metabolism, apoptosis, and metabolites transport. In plants, the expression of VDACs can be affected by different stresses, including drought, salinity and pathogen defense. In this study, we investigated the expression pattern of AtVDAC2 in A. thaliana and found ABA suppressed the accumulation of AtVDAC2 transcripts. Further, phenotype analysis of this VDAC deregulated-expression transgenic Arabidopsis plants indicated that AtVDAC2 anti-sense line showed an ABA-insensitivity phenotype during the early seedling development under ABA treatment. The results suggested that AtVDAC2 might be involved in ABA signaling in A. thaliana.