Evaluation of fifteen processing methods of hellgrammites based on the flavor characteristics.

To investigate suitable processing methods for improve the flavor while maintaining quality, hellgrammites were subjected to fifteen different processing methods. The samples were tested by sensory evaluation and were analyzed using HS-SPME-GC-MS. The sensory evaluation revealed that five methods for head and chest removal, three wine-fried methods, and three vinegar-roasting methods significantly reduced the levels of hexanal (3129.05 ± 45.77 µg/kg) and heptanal (436.72 ± 7.42 µg/kg), compounds responsible for fishy and earthy flavors, compared to raw samples. The latter two methods exhibited increased aroma flavor. PCA and OPLS-DA analyses suggested that acids, alcohols, and esters played a crucial role in flavor modification. Notably, vinegar-roasting methods demonstrated the highest acid content and had a substantial impact on volatile compounds. Additionally, boiling methods effectively reduced the levels of hazardous compounds, such as toluene and 1,3-Dimethyl-benzene. However, other methods did not exhibit similar efficacy in reducing hazardous compounds. The accumulation of hazardous compounds showed a decreasing trend in the whole insect, head removal, and head and chest removal groups. Moreover, the relative odor activity value consistently identified aldehyde compounds, including hexanal and heptanal, as the main contributors to aroma. Overall, boiling and head and chest removal procedures were suggested as precautionary measures during the initial processing of hellgrammites-based food products. The vinegar-roasting and wine-fried methods could be employed to impart desired flavors, aligning with consumers' preferences. These findings lay the foundation for standardizing processing techniques and ensuring the quality control of products derived from hellgrammites.

Unravel the regulatory mechanism of Yrr1p phosphorylation in response to vanillin stress in Saccharomyces cerevisiae.

Improving the resistance of Saccharomyces cerevisiae to vanillin, derived from lignin, will benefit the design of robust cell factories for lignocellulosic biorefining. The transcription factor Yrr1p mediates S. cerevisiae resistance to various compounds. In this study, eleven predicted phosphorylation sites were mutated, among which 4 mutants of Yrr1p, Y134A/E and T185A/E could improve vanillin resistance. Both dephosphorylated and phosphorylated mutations at Yrr1p 134 and 185 gathered in the nucleus regardless of the presence or absence of vanillin. However, the phosphorylated mutant Yrr1p inhibited target gene expression, while dephosphorylated mutants promoted expression. Transcriptomic analysis showed that the dephosphorylated Yrr1p T185 mutant, under vanillin stress, upregulated ribosome biogenesis and rRNA processing. These results demonstrate the mechanism by which Yrr1p phosphorylation regulates the expression of target genes. The identification of key phosphorylation sites in Yrr1p offers novel targets for the rational construction of Yrr1p mutants to improve resistance to other compounds.

Astaxanthin promotes mitochondrial biogenesis and antioxidant capacity in chronic high-intensity interval training.

PURPOSE: Reactive oxygen and nitrogen species are required for exercise-induced molecular adaptations; however, excessive exercise may cause cellular oxidative distress. We postulate that astaxanthin (ASX) can neutralize oxidative distress and stimulate mitochondrial biogenesis in high-intensity exercise-trained mice. METHODS: Six-week-old mice (n = 8/group) were treated with ASX (10 mg/kg BW) or placebo. Training groups participated in 30 min/day high-intensity interval training (HIIT) for 6 weeks. Gastrocnemius muscle was collected and assayed following the exercise training period. RESULTS: Compared to the HIIT control mice, the ASX-treated HIIT mice reduced malonaldehyde levels and upregulated the expression of Nrf2 and FOXO3a. Meanwhile, the genes NQO1 and GCLC, modulated by Nrf2, and SOD2, regulated by FOXO3a, and GPx4, were transcriptionally upregulated in the ASX-treated HIIT group. Meanwhile, the expression of energy sensors, AMPK, SIRT1, and SIRT3, increased in the ASX-treated HIIT group compared to the HIIT control group. Additionally, PGC-1α, regulated by AMPK and SIRT1, was upregulated in the ASX-treated HIIT group. Further, the increased PGC-1α stimulated the transcript of NRF1 and Tfam and mitochondrial proteins IDH2 and ATP50. Finally, the ASX-treated HIIT mice had upregulations in the transcript level of mitochondrial fusion factors, including Mfn1, Mfn2, and OPA1. However, the protein level of AMPK, SIRT1, and FOXO3a, and the transcript level of Nrf2, NQO1, PGC-1α, NRF1, Mfn1, Mfn2, and OPA1 decreased in the HIIT control group compared to the sedentary control group. CONCLUSION: Supplementation with ASX can reduce oxidative stress and promote antioxidant capacity and mitochondrial biogenesis during strenuous HIIT exercise in mice.

[Main components from cultivated and wild Nardostachyos Radix et Rhizoma by LC-MS and GC-MS].

In this study, ultra-performance liquid chromatography-quadrupole/time-of-flight mass spectrometry(UPLC-Q-TOF-MS) and gas chromatography-mass spectrometry(GC-MS) were combined with non-targeted metabonomic analysis based on multivariate statistics analysis, and the content of five indicative components in nardosinone was determined and compared by UPLC. The main chemical components of Nardostachyos Radix et Rhizoma with imitative wild cultivation and wild Nardostachyos Radix et Rhizoma were comprehensively analyzed. The results of multivariate statistical analysis based on liquid chromatography-mass spectrometry(LC-MS) and GC-MS were consistent. G1 and G2 of the imitative wild cultivation group and G8-G19 of the wild group were clustered into category 1, while G7 of the wild group and G3-G6 of the imitative wild cultivation group were clustered into category 2. After removing the outlier data of G1, G2, and G7, G3-G6 of the imitative wild cultivation group were clustered into one category, and G8-G19 of the wild group were clustered into the other category. Twenty-six chemical components were identified according to the positive and negative ion modes detected by LC-MS. The content of five indicative components(VIP>1.5) was determined using UPLC, revealing that chlorogenic acid, isochlorogenic acid A, isochlorogenic acid C, linarin, nardosinone, and total content in the imitative wild cultivation group were 1.85, 1.52, 1.26, 0.90, 2.93, and 2.56 times those in the wild group, respectively. OPLS-DA based on GC-MS obtained 10 diffe-rential peaks. Among them, the relative content of α-humulene and aristolene in the imitative wild cultivation group were extremely significantly(P<0.01) and significantly(P<0.05) higher than that in the wild group, while the relative content of 7 components such as 5,6-epoxy-3-hydroxy-7-megastigmen-9-one, γ-eudesmol, and juniper camphor and 12-isopropyl-1,5,9-trimethyl-4,8,13-cyclotetrade-catriene-1,3-diol was extremely significantly(P<0.01) and significantly(P<0.05) lower than that in the wild group, respectively. Therefore, the main chemical components of the imitative wild cultivation group and wild group were basically the same. However, the content of non-volatile components in the imitative wild cultivation group was higher than that in the wild group, and the content of some volatile components was opposite. This study provides scientific data for the comprehensive evaluation of the quality of Nardostachyos Radix et Rhizoma with imitative wild cultivation and wild Nardostachyos Radix et Rhizoma.

Combined transcriptomic and metabolomic analysis of Salmonella in the presence or absence of PhoP-PhoQ system under low Mg2+ conditions.

INTRODUCTION: Previous reports revealed the role played by Salmonella PhoP-PhoQ system in virulence activation, antimicrobial tolerance and intracellular survival, the impact of PhoP-PhoQ on cell metabolism has been less extensively described. OBJECTIVES: The aim of this study is to address whether and how the PhoP-PhoQ system affects the cell metabolism of Salmonella. METHODS: We constructed a Salmonella phoP deletion mutant strain TT-81 (PhoP-OFF), a Salmonella PhoP constitutively expressed strain TT-82 (PhoP-ON) and a wild-type Salmonella PhoP strain TT-80 (PhoP-N), using P22-mediated generalized transduction or λ Red-mediated targeted mutagenesis. We then measured the in vitro growth kinetics of all test strains and determined their metabolomic and transcriptomic profiles using gas chromatography coupled with tandem mass spectrometry (GC-MS/MS) and RNA-seq technique, respectively. RESULTS: Low-Mg2+ conditions impaired the growth of the phoP deletion mutant strain TT-81 (PhoP-OFF) dramatically. 42 metabolites in the wild-type PhoP strain TT-80 (PhoP-N) and 28 metabolites in the PhoP constitutively expressed strain TT-82 (PhoP-ON) changed by the absence of phoP. In contrast, the level of 19 compounds in TT-80 (PhoP-N) changed comparing to the PhoP constitutively expressed strain TT-82 (PhoP-N). The mRNA level of 95 genes in TT-80 (PhoP-N) changed when phoP was disrupted, wherein 78 genes downregulated and 17 genes upregulated. 106 genes were determined to be differentially expressed between TT-81 (PhoP-OFF) and TT-82 (PhoP-ON). While only 16 genes were found to differentially expressed between TT-82 (PhoP-ON) and TT-80 (PhoP-N). CONCLUSION: Our findings confirmed the impact of PhoP-PhoQ system on lipopolysaccharide (LPS) modification, energy metabolism, and the biosynthesis or transport of amino acids. Most importantly, we demonstrated that the turnover of a given metabolite could respond differentially to the level of phoP. Taken together, the present study provided new insights into the adaptation of Salmonella to the host environment and helped to characterize the impact of the PhoP-PhoQ system on the cell metabolism.

An actin-like protein PoARP9 involves in the regulation of development and cellulase and amylase expression in Penicillium oxalicum.

AIMS: In eukaryotic cells, chromatin remodelling complexes are essential for the accessibility of transcription factors to the specific regulating regions of downstream genes. Here, we identified an actin-like protein PoARP9 in cellulase production strain Penicillium oxalicum 114-2, which was an essential member of SWI/SNF complex. To investigate the physiological function of PoARP9 in transcriptional regulation, the coding gene Poarp9 was deleted in P. oxalicum 114-2. METHODS AND RESULTS: The absence of PoARP9 affected the colony growth on medium with glucose, cellulose or starch as sole carbon source. Meanwhile, the expression levels of major cellulase genes were all upregulated in ΔPoarp9 under the cellulase-inducing condition. In addition, the expression levels of amylase transcription activator AmyR as well as two major amylase genes were also increased in ΔPoarp9. CONCLUSIONS: These results demonstrated that chromatin remodelling affects the development and expression of cellulase and amylase in P. oxalicum. And the SWI/SNF complex member PoARP9 plays essential roles in these processes. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provided new insights into the regulation of cellulase and development in P. oxalicum. And the regulatory function of SWI/SNF complex member ARP9 towards cellulase and amylase expression in P. oxalicum was verified for the first time.

Chl1, an ATP-Dependent DNA Helicase, Inhibits DNA:RNA Hybrids Formation at DSB Sites to Maintain Genome Stability in S. pombe.

As an ATP-dependent DNA helicase, human ChlR1/DDX11 (Chl1 in yeast) can unwind both DNA:RNA and DNA:DNA substrates in vitro. Studies have demonstrated that ChlR1 plays a vital role in preserving genome stability by participating in DNA repair and sister chromatid cohesion, whereas the ways in which the biochemical features of ChlR1 function in DNA metabolism are not well understood. Here, we illustrate that Chl1 localizes to double-strand DNA break (DSB) sites and restrains DNA:RNA hybrid accumulation at these loci. Mutation of Chl1 strongly impairs DSB repair capacity by homologous recombination (HR) and nonhomologous end-joining (NHEJ) pathways, and deleting RNase H further reduces DNA repair efficiency, which indicates that the enzymatic activities of Chl1 are needed in Schizosaccharomyces pombe. In addition, we found that the Rpc37 subunit of RNA polymerase III (RNA Pol III) interacts directly with Chl1 and that deletion of Chl1 has no influence on the localization of Rpc37 at DSB site, implying the role of Rpc37 in the recruitment of Chl1 to this site.

[Comparison of odor and quality of Galli Gigerii Endothelium Corneum derived from domestic chickens and broilers].

Galli Gigerii Endothelium Corneum(GGEC) is commonly used for the clinical treatment of indigestion, vomiting, diarrhea, and infantile malnutrition with accumulation. In recent decades, omnivorous domestic chickens, the original source of GGEC, has been replaced by broilers, which may lead to significant changes in the quality of the yielding GGEC. Through subjective and objective sensory evaluation, biological evaluation, and chemical analysis, this study compared the odor and quality between GGEC derived from domestic chickens and that from broilers. The odor intensity between them was compared by odor profile analysis and it was found that the fishy odor of GGEC derived from domestic chickens was significantly weaker than that of GGEC from broilers. Headspace-solid phase microextraction-gas chromatography-triple quadrupole tandem mass spectrometry(HS-SPME/GC-QQQ-MS/MS) suggested that the overall odor-causing chemicals were consistent with the fishy odor-causing chemicals. According to the odor activity va-lue and the orthogonal partial least squares discriminant analysis(OPLS-DA) result, dimethyl trisulfide, 2-methoxy-3-isobutylpyrazine, and 2-methylisoborneol were responsible for the fishy odor(OAV≥1) and the content of fishy odor-causing chemicals in GGEC derived from broilers was 1.12-2.13 folds that in GGEC from domestic chickens. The average pepsin potency in GGEC derived from broilers was 15.679 U·mg~(-1), and the corresponding figure for the medicinal from domestic chickens was 26.529 U·mg~(-1). The results of pre-column derivatization reverse-phase high-performance liquid chromatography(RP-HPLC) assay showed that the content of total amino acids and digestion-promoting amino acids in domestic chickens-derived GGEC was 1.12 times and 1.15 times that in GGEC from broilers, and the bitter amino acid content was 1.21 times folds that of the latter. In conclusion, GGEC derived from domestic chickens had weaker fishy odor, stronger enzyme activity, higher content of digestion-promoting amino acids, and stronger bitter taste than GGEC from broilers. This study lays a scientific basis for studying the quality variation of GGEC and provides a method for identifying high-quality GGEC. Therefore, it is of great significance for the development and cultivation of GGEC as both food and medicine and breeding of corresponding varieties.

The Impact of SlyA on Cell Metabolism of Salmonella typhimurium: A Joint Study of Transcriptomics and Metabolomics.

SlyA is an important transcriptional regulator in Salmonella typhimurium (S. typhimurium). Numerous reports have indicated the impact of SlyA on the virulence of S. typhimurium. Less information regarding the role of SlyA in the cell metabolism of S. typhimurium is available. To close this gap, we compared the growth kinetics of an S. typhimurium wild-type strain to a slyA deletion mutant strain. The data suggested that the cell growth of S. typhimurium was impaired when slyA abolished, indicating that SlyA might affect the cell metabolism of S. typhimurium. To determine the role of SlyA in cell metabolism, we analyzed the metabolite profiles of S. typhimurium in the presence or absence of slyA using gas chromatography coupled with tandem mass spectrometry (GC-MS-MS). With the aim of appropriately interpreting the results obtained from metabolomics, a transcriptomic analysis on both the wild-type S. typhimurium and the slyA deletion mutant was performed. The metabolome data indicated that several glycolysis and lipid metabolism-associated pathways, including the turnover of glycerolipid, pyruvate, butanoate, and glycerophospholipid, were affected in the absence of slyA. In addition, the mRNA levels of several genes associated with glycolysis and lipid turnover were downregulated when slyA was deleted, including pagP, fadL, mgtB, iacp, and yciA. Collectively, these evidence suggested that SlyA affects the glycolysis and lipid turnover of S. typhimurium at a transcriptional level. The raw data of metabolomics is available in the MetaboLights database with an access number of MTBLS1858. The raw data of transcriptome is available in the Sequence Read Archive (SRA) database with an access number of PRJNA656165.

Quorum sensing-mediated protein degradation for dynamic metabolic pathway control in Saccharomyces cerevisiae.

Dynamic regulation has been widely applied to optimize metabolic flux distribution. However, compared with prokaryotes, quorum sensing-mediated pathway control is still very limited in Saccharomyces cerevisiae. In this study, we designed quorum sensing-regulated protein degradation circuits for dynamic metabolic pathway control in S. cerevisiae. The synthetic quorum sensing circuits were developed by integration of a plant hormone cytokinin system with the endogenous yeast Ypd1-Skn7 signal transduction pathway and the positive feedback circuits were optimized by promoter engineering. We then constructed an auxin-inducible protein degradation system and used quorum sensing circuits to regulate auxin synthesis to achieve dynamic control of protein degradation. As a demonstration, the circuits were applied to control Erg9 degradation to produce α-farnesene and the titer of α-farnesene increased by 80%. The population-regulated protein degradation system developed here extends dynamic regulation to the protein level in S. cerevisiae and is a promising approach for metabolic pathway control.

Proteomic analysis revealed the roles of YRR1 deletion in enhancing the vanillin resistance of Saccharomyces cerevisiae.

BACKGROUND: Vanillin is one of the important phenolic inhibitors in Saccharomyces cerevisiae for bioconversion of lignocellulosic materials and has been reported to inhibit the translation process in cells. In our previous studies, it was confirmed that the deletion of the transcription factor gene YRR1 enhanced vanillin resistance by promoting some translation-related processes at the transcription level. In this work, we investigated the effects of proteomic changes upon induction of vanillin stress and deletion of YRR1 to provide unique perspectives from a transcriptome analysis for comprehending the mechanisms of YRR1 deletion in the protective response of yeast to vanillin. RESULTS: In wild-type cells, vanillin reduced two dozens of ribosomal proteins contents while upregulated proteins involved in glycolysis, oxidative phosphorylation, and the pentose phosphate pathway in cells. The ratios of NADPH/NADP+ and NADH/NAD+ were increased when cells responded to vanillin stress. The differentially expressed proteins perturbed by YRR1 deletion were much more abundant than and showed no overlaps with transcriptome changes, indicating that Yrr1 affects the synthesis of certain proteins. Forty-eight of 112 upregulated proteins were involved in the stress response, translational and transcriptional regulation. YRR1 deletion increased the expression of HAA1-encoding transcriptional activator, TMA17-encoding proteasome assembly chaperone and MBF1-encoding coactivator at the protein level, as confirmed by ELISA. Cultivation data showed that the overexpression of HAA1 and TMA17 enhanced resistance to vanillin in S. cerevisiae. CONCLUSIONS: Cells conserve energy by decreasing the content of ribosomal proteins, producing more energy and NAD(P)H for survival in response to vanillin stress. Yrr1 improved vanillin resistance by increasing the protein quantities of Haa1, Tma17 and Mbf1. These results showed the response of S. cerevisiae to vanillin and how YRR1 deletion increases vanillin resistance at the protein level. These findings may advance our knowledge of how YRR1 deletion protects yeast from vanillin stress and offer novel targets for genetic engineering of designing inhibitor-resistant ethanologenic yeast strains.

Engineering transcription factor-based biosensors for repressive regulation through transcriptional deactivation design in Saccharomyces cerevisiae.

BACKGROUND: With the development of engineering the microbial cell factories, biosensors have been used widely for regulation of cellular metabolism and high-throughput screening. However, most of the biosensors constructed in Saccharomyces cerevisiae are designed for transcriptional activation. Very few studies have dedicated to the development of genetic circuit for repressive regulation, which is also indispensable for the dynamic control of metabolism. RESULTS: In this study, through transcriptional deactivation design, we developed transcription-factor-based biosensors to allow repressive regulation in response to ligand. Using a malonyl-CoA sensing system as an example, the biosensor was constructed and systematically engineered to optimize the dynamic range by comparing transcriptional activity of the activators, evaluating the positions and numbers of the operators in the promoter and comparing the effects of different promoters. A biosensor with 82% repression ratio was obtained. Based on this design principle, another two biosensors, which sense acyl-CoA or xylose and downregulate gene expression, were also successfully constructed. CONCLUSIONS: Our work systematically optimized the biosensors for repressive regulation in yeast for the first time. It provided useful framework to construct similar biosensors. Combining the widely reported biosensors for transcriptional activation with the biosensors developed here, it is now possible to construct biosensors with opposing transcriptional activities in yeast.

Significant association between DHFR promoter methylation and ischemic stroke in a Chinese hypertensive population.

OBJECTIVE: DHFR encodes dihydrofolate reductase, a major enzyme in the metabolism of folate, and is a candidate gene for ischemic stroke (IS). Therefore, we aimed to investigate the association between DHFR promoter methylation and IS in a Chinese population with primary hypertension. METHODS: Quantitative methylation-specific PCR was used to measure the level of DHFR promoter methylation. A multivariate logistic regression model was used to investigate the association between DHFR promoter methylation and IS. Receiver operating characteristic (ROC) curve analysis was used to evaluate the diagnostic value of DHFR promoter methylation for IS. RESULTS: The level of methylation of the DHFR promoter in the IS group was significantly lower than that in the hypertensive group (median [interquartile range]: 9.11 [2.81-16.20] vs 24.94 [7.16-56.45], P < .001). DHFR promoter methylation and homocysteine (Hcy) levels were both related to IS, with an ORs (95% CI) of 0.976 (0.967-0.984) and 1.057 (1.027-1.108), respectively. The areas under the curve for the diagnosis of DHFR promoter hypomethylation in IS were 0.603 (95% CI, 0.527-0.678) in men and 0.754 (95% CI, 0.693-0.815) in women. A dual-luciferase reporter assay revealed that the target sequence in the DHFR promoter upregulated gene expression. CONCLUSION: There is a significant association between methylation of the DHFR promoter and IS in this Chinese hypertensive population. Hypomethylation of the DHFR promoter may serve as a novel marker for the diagnosis of IS in women.

Increasing the level of 4-vinylguaiacol in top-fermented wheat beer by secretory expression of ferulic acid decarboxylase from Bacillus pumilus in brewer's yeast.

OBJECTIVE: The objective is to explore the effects of enhancing the activity of yeast ferulic acid decarboxylase (FDC1) on the level of 4-vinylguaiacol (4-VG) and the consumption of its precursor ferulic acid (FA) in top-fermented wheat beer. RESULTS: Expression of Bacillus pumilus FDC1 in brewer's yeast showed a better effect on the FDC1 activity than overexpression of the endogenous enzyme. The 4-VG content was increased by 34%, and the consumption time of FA was shortened from 48 to 12 h. Since the intracellular accumulation of the FDC1 substrate did not increase over time, to reduce the FA transport burden on cells and shorten the decarboxylation time, B. pumilus FDC1 was further secreted extracellularly. The resulted strain showed a 65% increase in 4-VG content in the FA-containing medium, and produced about 3 mg L-1 4-VG in the top-fermented wheat beer, increasing by 61% than control. However, further increasing the secretory expression level of FDC1 only accelerated FA consumption. CONCLUSIONS: These results suggested that appropriate secretion of bacterial FDC1 into wort could be used as a potential alternative strategy to increase the level of 4-VG in top-fermented wheat beer.

Development of novel surface display platforms for anchoring heterologous proteins in Saccharomyces cerevisiae.

BACKGROUND: Cell surface display of recombinant proteins has become a powerful tool for biotechnology and biomedical applications. As a model eukaryotic microorganism, Saccharomyces cerevisiae is an ideal candidate for surface display of heterologous proteins. However, the frequently used commercial yeast surface display system, the a-agglutinin anchor system, often results in low display efficiency. RESULTS: We initially reconstructed the a-agglutinin system by replacing two anchor proteins with one anchor protein. By directly fusing the target protein to the N-terminus of Aga1p and inserting a flexible linker, the display efficiency almost doubled, and the activity of reporter protein α-galactosidase increased by 39%. We also developed new surface display systems. Six glycosylphosphatidylinositol (GPI) anchored cell wall proteins were selected to construct the display systems. Among them, Dan4p and Sed1p showed higher display efficiency than the a-agglutinin anchor system. Linkers were also inserted to eliminate the effects of GPI fusion on the activity of the target protein. We further used the newly developed Aga1p, Dan4p systems and Sed1p system to display exoglucanase and a relatively large protein ß-glucosidase, and found that Aga1p and Dan4p were more suitable for immobilizing large proteins. CONCLUSION: Our study developed novel efficient yeast surface display systems, that will be attractive tools for biotechnological and biomedical applications.

Spatial Distribution of Organophosphorus and Brominated Flame Retardants in Surface Water, Sediment, Groundwater, and Wild Fish in Chengdu, China.

The occurrence and spatial distribution of 13 organophosphorus flame retardants (OPFRs), 11 polybrominated diphenyl ethers (PBDEs), and eight novel brominated flame retardants (NBFRs) were investigated in Jinjiang river water, sediment, crucian carp, and groundwater in Chengdu, China. OPFRs were predominant and ubiquitous contaminants in the Jinjiang river water, sediment, groundwater, fish muscle, fish gills, and viscera with concentrations ranging from 19.1 to 533 ng L-1, 12.5 to 253 ng g-1, 11.7 to 149 ng L-1, 114 to 2108 ng g-1 lipid weight (lw), 220 to 638 ng g-1 lw, and 116 to 1356 ng g-1 lw, respectively. The halogenated OPFRs were the primary pollutant in the Jinjiang river water samples, whereas nonhalogenated OPFRs were the dominant OPFRs in the sediments. Brominated flame retardants were not detected in the groundwater, whereas the NBFRs detected in aquatic environment at low frequency. The ΣPBDEs ranged from n.d. to 23.4 ng L-1 and n.d. to 48.7 ng g-1 in the Jinjiang river water and sediment, respectively. BDE-209 was dominant in the sediment samples with concentrations ranging from n.d. to 47.2 ng g-1. The PBDEs levels in the muscle, gills, and viscera of the crucian carp ranged from 10.6 to 90.6 ng g-1 lw, n.d. to 75.6 ng g-1 lw, and n.d. to 219 ng g-1 lw, respectively. BDE-47, chlorinated, and alkyl OPFRs were the main contaminants in the fish samples.

Developing synthetic hybrid promoters to increase constitutive or diauxic shift-induced expression in Saccharomyces cerevisiae.

Fine-tuning of the expression of genes is crucial for cell factory construction. Promoters are the most important tools to control gene expression. However, native promoters are often limited by their transcriptional ability. In this study, we sought to overcome the limitations of native promoters in Saccharomyces cerevisiae through the construction of hybrid promoter libraries for both constitutive promoters and promoters induced by diauxic shift. A series of hybrid constitutive promoters were constructed by combing the upstream activation sequences and changing the core promoter elements. The transcriptional capacity of the strongest promoter was 2-fold higher than that of the yeast native TEF1 promoter. Aside from the constitutive promoters, hybrid promoters that were induced in the post-diauxic phase were also constructed. These promoters had low transcriptional ability during growth on glucose and automatically activated upon growth with a diauxic shift. The strength of these promoters was also increased by replacing the core promoter with strong core promoters. Our study provides a series of constitutive and diauxic shift-induced promoters with a broad range of transcriptional capacity and will facilitate synthetic biology and metabolic engineering application.

Dry biorefining maximizes the potentials of simultaneous saccharification and co-fermentation for cellulosic ethanol production.

Despite the well-recognized merits of simultaneous saccharification and co-fermentation (SSCF) on relieving sugar product inhibition on cellulase activity, a practical concomitance difficulty of xylose with inhibitors in the pretreated lignocellulose feedstock prohibits the essential application of SSCF for cellulosic ethanol fermentation. To maximize the SSCF potentials for cellulosic ethanol production, a dry biorefining approach was proposed starting from dry acid pretreatment, disk milling, and biodetoxification of lignocellulose feedstock. The successful SSCF of the inhibitor free and xylose conserved lignocellulose feedstock after dry biorefining reached a record high ethanol titer at moderate cellulase usage and minimum wastewater generation. For wheat straw, 101.4 g/L of ethanol (equivalent to 12.8% in volumetric percentage) was produced with the overall yield of 74.8% from cellulose and xylose, in which the xylose conversion was 73.9%, at the moderate cellulase usage of 15 mg protein per gram cellulose. For corn stover, 85.1 g/L of ethanol (equivalent to 10.8% in volumetric percentage) is produced with the overall conversion of 84.7% from cellulose and xylose, in which the xylose conversion was 87.7%, at the minimum cellulase usage of 10 mg protein per gram cellulose. Most significantly, the SSCF operation achieved the high conversion efficiency by generating the minimum amount of wastewater. Both the fermentation efficiency and the wastewater generation in the current dry biorefining for cellulosic ethanol production are very close to that of corn ethanol production, indicating that the technical gap between cellulosic ethanol and corn ethanol has been gradually filled by the advancing biorefining technology.

Efficient yeast surface-display of novel complex synthetic cellulosomes.

BACKGROUND: The self-assembly of cellulosomes on the surface of yeast is a promising strategy for consolidated bioprocessing to convert cellulose into ethanol in one step. RESULTS: In this study, we developed a novel synthetic cellulosome that anchors to the endogenous yeast cell wall protein a-agglutinin through disulfide bonds. A synthetic scaffoldin ScafAGA3 was constructed using the repeated N-terminus of Aga1p and displayed on the yeast cell surface. Secreted cellulases were then fused with Aga2p to assemble the cellulosome. The display efficiency of the synthetic scaffoldin and the assembly efficiency of each enzyme were much higher than those of the most frequently constructed cellulosome using scaffoldin ScafCipA3 from Clostridium thermocellum. A complex cellulosome with two scaffoldins was also constructed using interactions between the displayed anchoring scaffoldin ScafAGA3 and scaffoldin I ScafCipA3 through disulfide bonds, and the assembly of secreted cellulases to ScafCipA3. The newly designed cellulosomes enabled yeast to directly ferment cellulose into ethanol. CONCLUSIONS: This is the first report on the development of complex multiple-component assembly system through disulfide bonds. This strategy could facilitate the construction of yeast cell factories to express synergistic enzymes for use in biotechnology.

Taxifolin prevents ß-amyloid-induced impairments of synaptic formation and deficits of memory via the inhibition of cytosolic phospholipase A2/prostaglandin E2 content.

Taxifolin is a potent flavonoid with anti-inflammatory activity. Taxifolin has been reported to decrease the accumulation of ß-amyloid (Aß), and reduce Aß-induced neurotoxicity. However, the detail molecular mechanism of taxifolin against Aß-induced neurotoxicity is largely unknown. In this study, we revealed the protective effects and the underlying mechanisms of taxifolin on the impairments of cognitive function and synapse formation induced by soluble Aß oligomers. Our results showed that taxifolin prevented neuronal cell death in a concentration-dependent manner. The recognition memory in novel object recognition tasks and the spatial memory in Morris water maze tests are significantly lower in the Alzheimer's disease (AD) model mice induced by hippocampal injection of Aß42. Taxifolin treatment prevented the recognitive and spatial memory deficits of the AD mice. 10 mg/kg taxifolin treatment also significantly prevented the decreased expression levels of PSD 95 induced by Aß42. Live cell imaging study showed that 2 h pre-treatment of taxifolin prevented the decrease in the number of filopodium and spine induced by Aß42 oligomers. Aß42 oligomers significantly increased the production of cytosolic phospholipase A2 (cPLA2), a crucial enzyme of pro-inflammatory mediator, and prostaglandin E2 (PGE2), a neuroinflammatory molecule. Taxifolin significantly reduced the content of cPLA2 and PGE2 induced by Aß42 both in the primary hippocampal neurons and hippocampal tissues. These results indicated that taxifolin might prevent Aß42 oligomer-induced synapse and cognitive impairments through decreasing cPLA2 and PGE2. Our study provided novel insights into the cellular mechanisms for the protective effects of taxifolin on AD.