Tianjin, China
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Li X.,Tianjin University | Li X.,SynBio Research Platform | Xu Q.-M.,Tianjin Normal University | Cheng J.-S.,Tianjin University | And 3 more authors.
Bioresource Technology | Year: 2016

The occurrence of sulfamethoxazole (SMX) in aquatic environment is a health concern. The presence of SMX significantly inhibited the laccase activity of Pycnoporus sanguineus with a lower removal efficiency of SMX. Although a laccase system with 2, 20-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) eliminated 100% SMX within 6 h, ABTS might cause an environmental issue. An alternative to SMX elimination is the coculture of Alcaligenes faecalis and P. sanguineus. The SMX removal efficiency at 48 h under the coculture with vitamins was higher than that under their pure culture alone, indicating that a coculture was more efficient in eliminating SMX than a pure culture. Only 1% SMX was detected in mycelia, indicating that SMX elimination is achieved primarily through biotransformation rather than adsorption. Laccase production by the coculture effectively inhibited the accumulations of N4-acetyl-SMX and N-hydroxy-SMX and alleviated the cytotoxicity of SMX transformation products. The mixture of SMX and sulfadiazine inhibited their removal efficiency. © 2016 Elsevier Ltd


Yang B.,Nankai University | Yang B.,Key Laboratory of Molecular Microbiology and Technology | Feng L.,Nankai University | Feng L.,Key Laboratory of Molecular Microbiology and Technology | And 7 more authors.
Nature Communications | Year: 2015

Enterohemorrhagic Escherichia coli (EHEC) is an important foodborne pathogen that infects humans by colonizing the large intestine. Here we identify a virulence-regulating pathway in which the biotin protein ligase BirA signals to the global regulator Fur, which in turn activates LEE (locus of enterocyte effacement) genes to promote EHEC adherence in the low-biotin large intestine. LEE genes are repressed in the high-biotin small intestine, thus preventing adherence and ensuring selective colonization of the large intestine. The presence of this pathway in all nine EHEC serotypes tested indicates that it is an important evolutionary strategy for EHEC. The pathway is incomplete in closely related small-intestinal enteropathogenic E. coli due to the lack of the Fur response to BirA. Mice fed with a biotin-rich diet show significantly reduced EHEC adherence, indicating that biotin might be useful to prevent EHEC infection in humans. © 2015 Macmillan Publishers Limited. All rights reserved.


Peng C.,Tianjin University | Luo H.,Tianjin University | Zhang X.,Tianjin University | Gao F.,Tianjin University | Gao F.,SynBio Research Platform
Frontiers in Microbiology | Year: 2015

DNA replication, one of the central events in the cell cycle, is the basis of biological inheritance. In order to be duplicated, a DNA double helix must be opened at defined sites, which are called DNA replication origins (ORIs). Unlike in bacteria, where replication initiates from a single replication origin, multiple origins are utilized in the eukaryotic genomes. Among them, the ORIs in budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe have been best characterized. In recent years, advances in DNA microarray and next-generation sequencing technologies have increased the number of yeast species involved in ORIs research dramatically. The ORIs in some non-conventional yeast species such as Kluyveromyces lactis and Pichia pastoris have also been genome-widely identified. Relevant databases of replication origins in yeast were constructed, then the comparative genomic analysis can be carried out. Here, we review several experimental approaches that have been used to map replication origins in yeast and some of the available web resources related to yeast ORIs. We also discuss the sequence characteristics and chromosome structures of ORIs in the four yeast species, which can be utilized to improve yeast replication origins prediction. © 2015 Peng, Luo, Zhang and Gao.


Niu X.,Tianjin University | Niu X.,SynBio Research Platform | Zhu Y.,Tianjin University | Zhu Y.,SynBio Research Platform | And 8 more authors.
Applied Microbiology and Biotechnology | Year: 2015

Butanol is highly toxic to cyanobacterial cells, which could restrict the future application of this renewable system in producing carbon neutral biofuel butanol. To seek knowledge regarding butanol tolerance in cyanobacteria, a library of response regulator (RR) gene mutants of Synechocystis sp. PCC 6803 was screened. The results showed that the deletion mutant of an orphan RR-encoding genes ll0039 was more sensitive to butanol than the wild type, while complementation of the ∆slr1037 mutant with the sll0039 gene recovered its tolerance to the same level as the wild type, suggesting that the sll0039 gene was involved in the regulation of tolerance against butanol. Further analysis employing an integrated liquid chromatography-mass spectrometry (LC-MS)-based and gas chromatography-mass spectrometry (GC-MS)-based metabolomics was conducted to determine the possible regulatory network of butanol tolerance mediated by Sll0039. LC-MS analysis allowed the identification of several metabolites, such as adenosine 5′-diphosphate (ADP)-glucose, dihydroxyacetone phosphate (DHAP), d-ribose 5-phosphate (R5P), d-glucose 6-phosphate (G6P), d-fructose 6-phosphate (F6P), α-ketoglutaric acid (AKG), uridine 5′-diphospho (UDP)-glucose, and nicotinamide adenine dinucleotide phosphate (NADP), which were differentially regulated between the wild type and the ∆sll0039 mutant grown under butanol stress, while GC-MS analysis identified 1, 2, and 2 metabolic modules associated with the sll0039 gene deletion at 24, 48, and 72 h, respectively, suggesting that they were under control directly or indirectly by Sll0039 RR. In addition, a metabolomic comparison of the metabolic responses to butanol stress was conducted in the ∆sll0039 mutant and the ∆slr1037 mutant previously found to be involved in butanol tolerance (Chen et al. Biotechnol Biofuels 7:89 2014a), and the results showed that the regulatory networks mediated by Sll0039 and Slr1037 could be functionally independent in Synechocystis. The results provided a metabolomic description of the butanol tolerance network regulated by Sll0039. © 2015, Springer-Verlag Berlin Heidelberg.


Luo H.,Tianjin University | Zhang C.-T.,Tianjin University | Gao F.,Tianjin University | Gao F.,SynBio Research Platform
Frontiers in Microbiology | Year: 2014

DNA replication is one of the most basic processes in all three domains of cellular life. With the advent of the post-genomic era, the increasing number of complete archaeal genomes has created an opportunity for exploration of the molecular mechanisms for initiating cellular DNA replication by in vivo experiments as well as in silico analysis. However, the location of replication origins (oriCs) in many sequenced archaeal genomes remains unknown. We present a web-based tool Ori-Finder 2 to predict oriCs in the archaeal genomes automatically, based on the integrated method comprising the analysis of base composition asymmetry using the Z-curve method, the distribution of origin recognition boxes identified by FIMO tool, and the occurrence of genes frequently close to oriCs. The web server is also able to analyze the unannotated genome sequences by integrating with gene prediction pipelines and BLAST software for gene identification and function annotation. The result of the predicted oriCs is displayed as an HTML table, which offers an intuitive way to browse the result in graphical and tabular form. The software presented here is accurate for the genomes with single oriC, but it does not necessarily find all the origins of replication for the genomes with multiple oriCs. Ori-Finder 2 aims to become a useful platform for the identification and analysis of oriCs in the archaeal genomes, which would provide insight into the replication mechanisms in archaea. © 2014 Luo, Zhang and Gao.


Qi Z.,Tianjin University | Qi Z.,SynBio Research Platform | Pei G.,Tianjin University | Pei G.,SynBio Research Platform | And 4 more authors.
Scientific Reports | Year: 2014

Microbial syntrophic metabolism has been well accepted as the heart of how methanogenic and other anaerobic microbial communities function. In this work, we applied a single-cell RT-qPCR approach to reveal gene-expression heterogeneity in a model syntrophic system of Desulfovibrio vulgaris and Methanosarcina barkeri, as compared with the D. vulgaris monoculture. Using the optimized primers and single-cell analytical protocol, we quantitatively determine gene-expression levels of 6 selected target genes in each of the 120 single cells of D. vulgaris isolated from its monoculture and dual-culture with M. barkeri. The results demonstrated very significant cell-to-cell gene-expression heterogeneity for the selected D. vulgaris genes in both the monoculture and the syntrophic dual-culture. Interestingly, no obvious increase in gene-expression heterogeneity for the selected genes was observed for the syntrophic dual-culture when compared with its monoculture, although the community structure and cell-cell interactions have become more complicated in the syntrophic dual-culture. In addition, the single-cell RT-qPCR analysis also provided further evidence that the gene cluster (DVU0148-DVU0150) may be involved syntrophic metabolism between D. vulgaris and M. barkeri. Finally, the study validated that single-cell RT-qPCR analysis could be a valuable tool in deciphering gene functions and metabolism in mixed-cultured microbial communities.


Wang Y.,Tianjin University | Wang Y.,SynBio Research Platform | Chen L.,Tianjin University | Chen L.,SynBio Research Platform | And 2 more authors.
Biotechnology for Biofuels | Year: 2016

Background: 3-hydroxypropionic acid (3-HP) is an important platform chemical with a wide range of applications. In our previous study, the biosynthetic pathway of 3-HP was constructed and optimized in cyanobacterium Synechocystis sp. PCC 6803, which led to 3-HP production directly from CO2 at a level of 837.18 mg L−1 (348.8 mg/g dry cell weight). As the production and accumulation of 3-HP in cells affect cellular metabolism, a better understanding of cellular responses to 3-HP synthesized internally in Synechocystis will be important for further increasing 3-HP productivity in cyanobacterial chassis. Results: Using a engineered 3-HP-producing SM strain, in this study, the cellular responses to 3-HP internally produced were first determined using a quantitative iTRAQ-LC-MS/MS proteomics approach and a LC-MS-based targeted metabolomics. A total of 2264 unique proteins were identified, which represented about 63 % of all predicted protein in Synechocystis in the proteomic analysis; meanwhile intracellular abundance of 24 key metabolites was determined by a comparative metabolomic analysis of the 3-HP-producing strain SM and wild type. Among all identified proteins, 204 proteins were found up-regulated and 123 proteins were found down-regulated, respectively. The proteins related to oxidative phosphorylation, photosynthesis, ribosome, central carbon metabolism, two-component systems and ABC-type transporters were up-regulated, along with the abundance of 14 metabolites related to central metabolism. The results suggested that the supply of ATP and NADPH was increased significantly, and the precursor malonyl-CoA and acetyl-CoA may also be supplemented when 3-HP was produced at a high level in Synechocystis. Confirmation of proteomic and metabolomic results with RT-qPCR and gene-overexpression strains of selected genes was also conducted, and the overexpression of three transporter genes putatively involved in cobalt/nickel, manganese and phosphate transporting (i.e., sll0385, sll1598 and sll0679) could lead to an increased 3-HP production in Synechocystis. Conclusions: The integrative analysis of up-regulated proteome and metabolome data showed that to ensure the high-efficient production of 3-HP and the normal growth of Synechocystis, multiple aspects of cells metabolism including energy, reducing power supply, central carbon metabolism, the stress responses and protein synthesis were enhanced in Synechocystis. The study provides an important basis for further engineering cyanobacteria for high 3-HP production. © 2016 The Author(s).


Lin F.,Tianjin University | Guo X.,Tianjin University | Lu W.,Tianjin University | Lu W.,SynBio Research Platform
Antonie van Leeuwenhoek, International Journal of General and Molecular Microbiology | Year: 2015

Ginsenosides are the major pharmacological components in ginseng. Microorganisms from a ginseng field were isolated to identify transformation of ginsenosides. Based on HPLC and LC–MS analysis, strain LFJ1403 showed strong activities to transform ginsenoside Rb1 to Rd as the sole product. Phylogenetic analysis of 18S rDNA indicated that LFJ1403 belonged to Aspergillus versicolor. Through comparing four systems of transforming Rb1 to Rd, strain LFJ1403 was found to secrete ginsenoside-converting enzymes in the spore production phase of plate culture. This result suggested that the enzyme could be directly obtained from the plate. The spore suspension, which contained the exocrine enzyme, was easy to prepare and efficient for biotransformation of ginsenoside Rb1 to Rd. Further study showed that the maximum bioconversion rate was 96 % (w/w) in shake flasks when a spore suspension system was used with optimized biotransformation conditions. Scale-up of this system to 2L resulted in an 85 % conversion rate. The ginsenoside Rb1 converting enzyme was separated by gradient HPLC with Q-Sepharose column, and its β-glucosidase activity and Rb1-converting ability was assayed by the 4-Nitrophenyl-β-d-glucopyranoside (PNPG) method and HPLC with C18 column, respectively. We obtained 130 U ml−1 enzymatic activity with the purified β-glucosidase. This is the first report on efficiently converting ginsenoside using extracellular enzyme directly from the fungus spore production phase of solid culture. © 2015, Springer International Publishing Switzerland.


Wang G.,Tianjin University | Wang G.,SynBio Research Platform | Huang D.,Nankai University | Li Y.,Tianjin University | And 5 more authors.
Bioresource Technology | Year: 2015

In this work, wheat bran (WB) was utilized as feedstock to synthesize fumaric acid by Rhizopus oryzae. Firstly, the pretreatment process of WB by dilute sulfuric acid hydrolysis undertaken at 100. °C for 30. min offered the best performance for fumaric acid production. Subsequently, through optimizing the seed culture medium, a suitable morphology (0.55. mm pellets diameter) of R. oryzae was obtained. Furthermore, a metabolic-based approach was developed to profile the differences of intracellular metabolites concentration of R. oryzae between xylose (the abundant sugar in wheat bran hydrolysate (WBH)) and glucose metabolism. The xylitol, sedoheptulose 7-phosphate, ribulose 5-phosphate, glucose 6-phosphate, proline and serine were responsible for fumaric acid biosynthesis limitation in xylose fermentation. Consequently, regulation strategies were proposed, leading to a 149% increase in titer (up to 15.4. g/L). Finally, by combinatorial regulation strategies the highest production was 20.2. g/L from WBH, 477% higher than that of initial medium. © 2014 Elsevier Ltd.


Liang D.-M.,Tianjin University | Liang D.-M.,Key Laboratory of Systems Bioengineering | Liang D.-M.,SynBio Research Platform | Liu J.-H.,Tianjin University | And 14 more authors.
Chemical Society Reviews | Year: 2015

Glycosylation reactions mainly catalyzed by glycosyltransferases (Gts) occur almost everywhere in the biosphere, and always play crucial roles in vital processes. In order to understand the full potential of Gts, the chemical and structural glycosylation mechanisms are systematically summarized in this review, including some new outlooks in inverting/retaining mechanisms and the overview of GT-C superfamily proteins as a novel Gt fold. Some special features of glycosylation and the evolutionary studies on Gts are also discussed to help us better understand the function and application potential of Gts. Natural product (NP) glycosylation and related Gts which play important roles in new drug development are emphasized in this paper. The recent advances in the glycosylation pattern (particularly the rare C- and S-glycosylation), reversibility, iterative catalysis and protein auxiliary of NP Gts are all summed up comprehensively. This review also presents the application of NP Gts and associated studies on synthetic biology, which may further broaden the mind and bring wider application prospects. © The Royal Society of Chemistry.

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