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1.
Objective
To construct a promoter probe vector, pBE-bgaB, to screen strong promoters from Bacillus amyloliquefaciens.Results
266 colonies containing active promoter elements from the genomic DNA of B. amyloliquefaciens were identified. Among these, promoter P41 exhibited the strongest β-Gal activity in Escherichia coli and B. amyloliquefaciens. Sequence analysis showed that promoter P41 contained P ykuN , a ykuN gene encoding flavodoxin. Optimization of the ribosome-binding site from P41 to P382 improved β-Gal activity by ~ 200%.Conclusion
A new strong promoter for protein expression and genetic engineering of Bacillus species.2.
Lifei Chen Chunling Ma Ruiming Wang Jianlou Yang Haijie Zheng 《Biotechnology letters》2016,38(10):1769-1774
Objectives
To improve 1,3-propanediol (1,3-PD) production and reduce byproduct concentration during the fermentation of Klebsiella pneumonia.Results
Klebsiella. pneumonia 2-1ΔldhA, K. pneumonia 2-1ΔaldH and K. pneumonia 2-1ΔldhAΔaldH mutant strains were obtained through deletion of the ldhA gene encoding lactate dehydrogenase required for lactate synthesis and the aldH gene encoding acetaldehyde dehydrogenase involved in the synthesis of ethanol. After fed-batch fermentation, the production of 1,3-PD from glycerol was enhanced and the concentrations of byproducts were reduced compared with the original strain K. pneumonia 2-1. The maximum yields of 1,3-PD were 85.7, 82.5 and 87.5 g/l in the respective mutant strains.Conclusion
Deletion of either aldH or ldhA promoted 1,3-PD production in K. pneumonia.3.
Background
The ability to respond rapidly to fluctuations in environmental changes is decisive for cell survival. Under these conditions trehalose has an essential protective function and its concentration increases in response to enhanced expression of trehalose synthase genes, TPS1, TPS2, TPS3 and TSL1. Intriguingly, the NTH1 gene, which encodes neutral trehalase, is highly expressed at the same time. We have previously shown that trehalase remains in its inactive non-phosphorylated form by the action of an endogenous inhibitor. Recently, a comprehensive two-hybrid analysis revealed a 41-kDa protein encoded by the YLR270w ORF, which interacts with NTH1p.Results
In this work we investigate the correlation of this Trehalase Associated Protein, in trehalase activity regulation. The neutral trehalase activity in the ylr270w mutant strain was about 4-fold higher than in the control strain. After in vitro activation by PKA the ylr270w mutant total trehalase activity increased 3-fold when compared to a control strain. The expression of the NTH1 gene promoter fused to the heterologous reporter lacZ gene was evaluated. The mutant strain lacking YLR270w exhibited a 2-fold increase in the NTH1-lacZ basal expression when compared to the wild type strain.Conclusions
These results strongly indicate a central role for Ylr270p in inhibiting trehalase activity, as well as in the regulation of its expression preventing a wasteful futile cycle of synthesis-degradation of trehalose.4.
Elaheh Sajadi Valiollah Babaipour Ali Asghar Deldar Bagher Yakhchali Seyed Safa-Ali Fatemi 《Biotechnology letters》2017,39(9):1395-1401
Objectives
To evaluate the crystallinity index of the cellulose produced by Escherichia coli Nissle 1917 after heterologous expression of the cellulose synthase subunit D (bcsD) gene of Gluconacetobacter xylinus BPR2001.Results
The bcsD gene of G. xylinus BPR2001 was expressed in E. coli and its protein product was visualized using SDS-PAGE. FTIR analysis showed that the crystallinity index of the cellulose produced by the recombinants was 0.84, which is 17% more than that of the wild type strain. The increased crystallinity index was also confirmed by X-ray diffraction analysis. The cellulose content was not changed significantly after over-expressing the bcsD.Conclusion
The bcsD gene can improve the crystalline structure of the bacterial cellulose but there is not any significant difference between the amounts of cellulose produced by the recombinant and wild type E. coli Nissle 1917.5.
Objectives
To characterize the genes responsible for ethanol utilization in Pichia pastoris.Results
ADH3 (XM_002491337) and ADH (FN392323) genes were disrupted in P. pastoris. The ADH3 mutant strain, MK115 (Δadh3), lost its ability to grow on minimal ethanol media but produced ethanol in minimal glucose medium. ADH3p was responsible for 92 % of total Adh enzyme activity in glucose media. The double knockout strain MK117 (Δadh3Δadh) also produced ethanol. The Adh activities of X33 and MK116 (Δadh) strains were not different. Thus, the ADH gene does not play a role in ethanol metabolism.Conclusion
The PpADH3 is the only gene responsible for consumption of ethanol in P. pastoris.6.
Objectives
To reduce the amount of citrulline produced by arginine-consuming bacteria in the moromi mash during soy sauce production.Results
Bacillus amyloliquefaciens JY06, a salt-tolerant strain with high arginine consumption ability and low citrulline accumulation capacity, was isolated from moromi mash. The concentration of citrulline was decreased from 26.8 to 5.1 mM and ethyl carbamate in soy sauce, after sterilization, decreased from 97 to 17 μg kg?1 when B. amyloliquefaciens JY06 was added during fermentation. The aroma of the sauce was improved by increasing the ester content.Conclusions
B. amyloliquefaciens JY06 is a beneficial bacterium that can be used in soy sauce fermentation to eliminate ethyl carbonate and enhance the flavor of the sauce.7.
Mathivanan Ashokkumar Geetha Irudayaraj Nandakumar Yellapu Arulsamy Mary Manonmani 《World journal of microbiology & biotechnology》2018,34(8):116
A strain of Bacillus amyloliquefaciens (VCRC B483) exhibiting mosquito pupicidal, keratinase and antimicrobial activities was isolated from mangrove forest ecosystem of Andaman and Nicobar Islands. Molecular characterization of the strain showed the presence of lipopeptide encoding bmyC gene. Phylogenetic tree based on protein sequence of this gene exhibited homology with mycosubtilin synthetase of Bacilus atropheus and Iturin synthetase of Bacillus subtilis and B. amyloliquefaciens. This is the first report on the evolutionary conservation of amino acids concerned with the function and structure of bmyC protein of B. amyloliquefaciens. The presence of valine at the 1197th position in our strain was found to be unique and different from the existing strains of B. subtilis and B. amyloliquefaciens. Molecular modelling studies revealed significant changes in the structure of epimerization domain of the bmyC protein with A1197V variation. Crude metabolite of this strain exhibited antifungal activity against Fusarium sp. and Carvularia sp. 相似文献
8.
Marie GB Hansen Mette Christoffersen Line R Thuesen Morten R Petersen Anders M Bojesen 《Acta veterinaria Scandinavica》2010,52(1):3
Background
Borrelia burgdorferi sensu lato and Anaplasma phagocytophilum are able to infect horses. However, the extend to which Danish horses are infected and seroconvert due to these two bacteria is unknown. The aim of the present study was to evaluate the seroprevalence of B. burgdorferi sensu lato and A. phagocytophilum in Danish horses.Methods
A total of 390 blood samples collected from all major regions of Denmark and with a geographical distribution corresponding to the density of the Danish horse population were analyzed. All samples were examined for the presence of antibodies against B. burgdorferi sensu lato and A. phagocytophilum by the use of the SNAP®4DX ® ELISA test.Results
Overall, 29.0% of the horses were seropositive for B. burgdorferi sensu lato whereas 22.3% were seropositive for A. phagocytophilum.Conclusions
Antibodies against B burgdorferi sensu lato and A. phagocytophilum are commonly found among Danish horses thus showing that Danish horses are frequently infected by these organisms.9.
Xuechang Wu Lijie Zhang Xinna Jin Yahong Fang Ke Zhang Lei Qi Daoqiong Zheng 《Biotechnology letters》2016,38(7):1097-1106
10.
Objectives
To enhance acid tolerance of Candida glabrata for pyruvate production by engineering AMP metabolism.Results
The physiological function of AMP deaminase in AMP metabolism from C. glabrata was investigated by deleting or overexpresseing the corresponding gene, CgAMD1. At pH 4, CgAMD1 overexpression resulted in 59 and 51% increases in biomass and cell viability compared to those of wild type strain, respectively. In addition, the intracellular ATP level of strain Cgamd1Δ/CgAMD1 was down-regulated by 22%, which led to a 94% increase in pyruvate production. Further, various strengths of CgAMD1 expression cassettes were designed, thus resulting in a 59% increase in pyruvate production at pH 4. Strain Cgamd1Δ/CgAMD1 (H) was grown in a 30 l batch bioreactor at pH 4, and pyruvate reached 46.1 g/l.Conclusion
CgAMD1 overexpression plays an active role in improving acid tolerance and pyruvate fermentation performance of C. glabrata at pH 4.11.
Hyun-Soo Kim 《Biotechnology letters》2016,38(11):1955-1960
Objective
To identify a novel gene responsible for organic solvent-tolerance by screening a transposon-mediated deletion mutant library based on Saccharomyces cerevisiae L3262.Results
One strain tolerant of up to 0.5 % (v/v) n-hexane and cyclohexane was isolated. The determination of transposon insertion site identified one gene, YLR162W, and revealed disruption of the ORF of this gene, indicating that organic solvent tolerance can be conferred. Such a tolerant phenotype reverted to the sensitive phenotype on the autologous or overexpression of this gene. This transposon mutant grew faster than the control strain when cultured at 30 °C in YPD medium containing 0.5 % (v/v) n-hexane and cyclohexane respectively.Conclusion
Disruption of YLR162W in S. cerevisiae results in increased tolerance to organic solvents.12.
Young-Saeng Kim Il-Sup Kim Joseph S. Boyd Arnaud Taton James W. Golden Ho-Sung Yoon 《Biotechnology letters》2017,39(10):1499-1507
Objectives
To improve the oxidative stress tolerance, biomass yield, and ascorbate/dehydroascorbate (AsA/DHA) ratio of Synechococcus elongatus PCC 7942 in the presence of H2O2, by heterologous expression of the dehydroascorbate reductase (DHAR) gene from Brassica juncea (BrDHAR).Results
Under H2O2 stress, overexpression of BrDHAR in the transgenic strain (BrD) of S. elongatus greatly increased the AsA/DHA ratio. As part of the AsA recycling system, the oxidative stress response induced by reactive oxygen species was enhanced, and intracellular H2O2 level decreased. In addition, under H2O2 stress conditions, the BrD strain displayed increased growth rate and biomass, as well as higher chlorophyll content and deeper pigmentation than did wild-type and control strains.Conclusion
By maintaining the AsA pool and redox homeostasis, the heterologous expression of BrDHAR increased S. elongatus tolerance to H2O2 stress, improving the biomass yield under these conditions. The results suggest that the BrD strain of S. elongatus, with its ability to attenuate the deleterious effects of ROS caused by environmental stressors, could be a promising platform for the generation of biofuels and other valuable bioproducts.13.
14.
Jiwei Mao Quanli Liu Xiaofei Song Hesuiyuan Wang Hui Feng Haijin Xu Mingqiang Qiao 《Biotechnology letters》2017,39(7):977-982
Objective
To identify new enzymatic bottlenecks of l-tyrosine pathway for further improving the production of l-tyrosine and its derivatives.Result
When ARO4 and ARO7 were deregulated by their feedback resistant derivatives in the host strains, the ARO2 and TYR1 genes, coding for chorismate synthase and prephenate dehydrogenase were further identified as new important rate-limiting steps. The yield of p-coumaric acid in the feedback-resistant strain overexpressing ARO2 or TYR1, was significantly increased from 6.4 to 16.2 and 15.3 mg l?1, respectively. Subsequently, we improved the strain by combinatorial engineering of pathway genes increasing the yield of p-coumaric acid by 12.5-fold (from 1.7 to 21.3 mg l?1) compared with the wild-type strain. Batch cultivations revealed that p-coumaric acid production was correlated with cell growth, and the formation of by-product acetate of the best producer NK-M6 increased to 31.1 mM whereas only 19.1 mM acetate was accumulated by the wild-type strain.Conclusion
Combinatorial metabolic engineering provides a new strategy for further improvement of l-tyrosine or other metabolic biosynthesis pathways in S. cerevisiae.15.
Wenyi Sun Xiaobing Yang Xueying Wang Xinping Lin Yanan Wang Sufang Zhang Yushi Luan Zongbao K. Zhao 《Biotechnology letters》2017,39(7):1001-1007
Objectives
To target a carotenoid biosynthetic gene in the oleaginous yeast Rhodosporidium toruloides by using the Agrobacterium-mediated transformation (AMT) method.Results
The RHTO_04602 locus of R. toruloides NP11, previously assigned to code the carotenoid biosynthetic gene CRTI, was amplified from genomic DNA and cloned into the binary plasmid pZPK-mcs, resulting in pZPK-CRT. A HYG-expression cassette was inserted into the CRTI sequence of pZPK-CRT by utilizing the restriction-free clone strategy. The resulted plasmid was used to transform R. toruloides cells according to the AMT method, leading to a few white transformants. Sequencing analysis of those transformants confirmed homologous recombination and insertional inactivation of CRTI. When the white variants were transformed with a CRTI-expression cassette, cells became red and produced carotenoids as did the wild-type strain NP11.Conclusions
Successful homologous targeting of the CrtI locus confirmed the function of RHTO_04602 in carotenoids biosynthesis in R. toruloides. It provided valuable information for metabolic engineering of this non-model yeast species.16.
Objectives
To deregulate the purine operon of the purine biosynthetic pathway and optimize energy generation of the respiratory chain to improve the yield of guanosine in Bacillus amyloliquefaciens XH7.Results
The 5′-untranslated region of the purine operon, which contains the guanine-sensing riboswitch, was disrupted. The native promoter Pw in B. amyloliquefaciens XH7 was replaced by different strong promoters. Among the promoter replacement mutants, XH7purE::P41 gave the highest guanosine yield (16.3 g/l), with an increase of 23% compared with B. amyloliquefaciens XH7. The relative expression levels of the purine operon genes (purE, purF, and purD) in the XH7purE::P41 mutant were upregulated. The concentration of inosine monophosphate (IMP), the primary intermediate in the purine pathway, was also significantly increased in the XH7purE::P41 mutant. Combined modification of the low-coupling branched respiratory chains (cytochrome bd oxidase) improved guanosine production synergistically. The final guanosine yield in the XH7purE::P41△cyd mutant increased by 41% to 19 g/l compared with B. amyloliquefaciens XH7.Conclusion
The combined modification strategy used in this study is a novel approach to improve the production of guanosine in industrial bacterial strains.17.
Xue-Ting Chen Jun-Bin Ji Yong-Chuang Liu Bin Ye Chao-Yang Zhou Xin Yan 《Biotechnology letters》2016,38(12):2109-2117
Objectives
To induce natural genetic competence in Bacillus amyloliquefaciens isolates through overexpression of the master regulator, ComK, from B. subtilis (ComK Bsu ).Results
Plasmid pUBXC carrying the xylose-inducible comK expression cassette was constructed using plasmid pUB110 as a backbone. Plasmid pUBXC could be transferred from B. subtilis to B. amyloliquefaciens through plasmid pLS20-mediated biparental conjugation. After being induced by xylose, four B. amyloliquefaciens strains harbouring plasmid pUBXC developed genetic competence. Under optimal conditions, the transformation efficiencies of plasmid DNA ranged from 129 ± 20.6 to 1.7 ± 0.1 × 105 cfu (colony-forming units) per μg DNA, and the transformation efficiencies of PCR-assembled deletion constructs ranged from 3.2 ± 0.76 to 3.5 ± 0.42 × 104 cfu per μg DNA in the four tested strains.Conclusion
Artificial induction of genetic competence through overexpressing ComK Bsu in B. amyloliquefaciens completed the tasks of replicative plasmid delivery and gene knockout via direct transformation of PCR-generated deletion cassettes.18.
Pegah Amiri Azar Shahpiri Mohammad Ali Asadollahi Fariborz Momenbeik Siavash Partow 《Biotechnology letters》2016,38(3):503-508
Objectives
To engineer the yeast Saccharomyces cerevisiae for the heterologous production of linalool.Results
Expression of linalool synthase gene from Lavandula angustifolia enabled heterologous production of linalool in S. cerevisiae. Downregulation of ERG9 gene, that encodes squalene synthase, by replacing its native promoter with the repressible MET3 promoter in the presence of methionine resulted in accumulation of 78 µg linalool l?1 in the culture medium. This was more than twice that produced by the control strain. The highest linalool titer was obtained by combined repression of ERG9 and overexpression of tHMG1. The yeast strain harboring both modifications produced 95 μg linalool l?1.Conclusions
Although overexpression of tHMG1 and downregulation of ERG9 enhanced linalool titers threefold in the engineered yeast strain, alleviating linalool toxicity is necessary for further improvement of linalool biosynthesis in yeast.19.
Sen Miao Hao Wu Yue Zhao Qinggele Caiyin Yanni Li Jianjun Qiao 《Biotechnology letters》2018,40(6):941-948
20.
Jinxiang Zhu Qiaoyun Zhu Ruiqing Gong Qin Xu Menghao Cai Tianyi Jiang Xiangshan Zhou Mian Zhou Yuanxing Zhang 《Biotechnology letters》2018,40(9-10):1365-1376