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1.
Objective
To improve the production of trans-10,cis-12-conjugated linoleic acid (t10,c12-CLA) from linoleic acid in recombinant Yarrowia lipolytica.Results
Cells of the yeast were permeabilized by freeze/thawing. The optimal conditions for t10,c12-CLA production by the permeabilized cells were at 28 °C, pH 7, 200 rpm with 1.5 g sodium acetate l?1, 100 g wet cells l?1, and 25 g LA l?1. Under these conditions, the permeabilized cells produced 15.6 g t10,c12-CLA l?1 after 40 h, with a conversion yield of 62 %. The permeabilized cells could be used repeatedly for three cycles, with the t10,c12-CLA extracellular production remaining above 10 g l?1.Conclusion
Synthesis of t10,c12-CLA was achieved using a novel method, and the production reported in this work is the highest value reported to date.2.
Cheng-Hua Wang Tong-Xin Zhao Mei Li Chong Zhang Xin-Hui Xing 《Biotechnology letters》2016,38(2):337-344
Objective
To characterize a novel xanthine dehydrogenase (XDH) from Acinetobacter baumannii by recombinant expression in Escherichia coli and to assess its potential for industrial applications.Results
The XDH gene cluster was cloned from A. baumannii CICC 10254, expressed heterologously in E. coli and purified to homogeneity. The purified recombinant XDH consisted of two subunits with the respective molecular weights of 87 kDa and 56 kDa according to SDS-PAGE. XDH catalysis was optimum at pH 8.5 and 40–45 °C, was stable under alkaline conditions (pH 7–11) and the half-inactivation temperature was 60 °C. The K m, turnover number and catalytic efficiency for xanthine were 25 μM, 69 s?1 and 2.7 μM?1 s?1, respectively, which is an improvement over XDHs characterized previously. A. baumannii XDH is less than 50 % identical to previously identified XDH orthologs from other species, and is the first from the Acinetobacter genus to be characterized.Conclusion
The novel A. baumannii enzyme was found to be among the most active, thermostable and alkaline-tolerant XDH enzymes reported to date and has potential for use in industrial applications.3.
4.
Objective
To investigate green synthesis of gold nanoparticles (AuNPs) by Trichosporon montevideense, and to study their reduction of nitroaromatics.Results
AuNPs had a characteristic absorption maximum at 535 nm. Scanning electron microscopy images revealed that the biosynthesized nanoparticles were attached on the cell surface. X-ray diffraction analysis indicated that the particles formed as face-centered cubic (111)-oriented crystals. The average size of AuNPs decreased from 53 to 12 nm with increasing biomass concentration. The catalytic reduction of 2-nitrophenol, 3-nitrophenol, 4-nitrophenol, o-nitrophenylamine and m-nitrophenylamine (0.1 mM) by NaBH4 had reaction rate constants of 0.32, 0.44, 0.09, 0.24 and 0.39 min?1 with addition of 1.45 × 10?2 mM AuNPs.Conclusions
An eco-friendly approach for synthesis of AuNPs by T. montevideense is reported for the first time. The biogenic AuNPs could serve as efficient catalysts for hydrogenation of various nitroaromatics.5.
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.6.
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.7.
Deqiang Zhu Jianrong Wu Xiaobei Zhan Li Zhu Zhiyong Zheng Minjie Gao 《Biotechnology letters》2017,39(2):227-234
Objectives
N-Acetyl-d-neuraminic acid (Neu5Ac) is often synthesized from exogenous N-acetylglucosamine (GlcNAc) and excess pyruvate. We have previously constructed a recombinant Escherichia coli strain for Neu5Ac production using GlcNAc and intracellular phosphoenolpyruvate (PEP) as substrates (Zhu et al. Biotechnol Lett 38:1–9, 2016).Results
PEP synthesis-related genes, pck and ppsA, were overexpressed within different modes to construct PEP-supply modules, and their effects on Neu5Ac production were investigated. All the PEP-supply modules enhanced Neu5Ac production. For the best module, pCDF-pck-ppsA increased Neu5Ac production to 8.6 ± 0.15 g l?1, compared with 3.6 ± 0.15 g l?1 of the original strain. Neu5Ac production was further increased to 15 ± 0.33 g l?1 in a 1 l fermenter.Conclusions
The PEP-supply module can improve the intracellular PEP supply and enhance Neu5Ac production, which benefited industrial Neu5Ac production.8.
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.9.
Ryosuke Fujiwara Shuhei Noda Yoshifumi Kawai Tsutomu Tanaka Akihiko Kondo 《Biotechnology letters》2016,38(9):1543-1549
Objectives
To find a novel host for the production of 4-vinylphenol (4VPh) by screening Streptomyces species.Results
The conversion of p-coumaric acid (pHCA) to 4VPh in Streptomyces mobaraense was evaluated using a medium containing pHCA. S. mobaraense readily assimilated pHCA after 24 h of cultivation to produce 4VPh. A phenolic acid decarboxylase, derived from S. mobaraense (SmPAD), was purified following heterologous expression in Escherichia coli. SmPAD was evaluated under various conditions, and the enzyme’s kcat/Km value was 0.54 mM ?1 s?1. Using intergenetic conjugation, a gene from Rhodobacter sphaeroides encoding a tyrosine ammonia lyase, which catalyzes the conversion of l-tyrosine to p-coumaric acid, was introduced into S. mobaraense. The resulting S. mobaraense transformant produced 273 mg 4VPh l?1 from 10 g glucose l?1.Conclusion
A novel strain suitable for the production of 4VPh and potentially other aromatic compounds was isolated.10.
Objective
To ascertain the effect of chitin-binding domain (ChBD) and fibronectin type III domain (FN3) on the characterization of the intact chitinase from Bacillus thuringiensis.Results
An intact chitinase gene (chi74) from B. thuringiensis HZP7 and its truncated genes (chi54, chi63 and chi66) were expressed in Escherichia coli BL21. The expression products were analyzed after purification. All chitinases were active from pH 4–7.5 and from 20 to 80 °C with identical optimal: pH 5.5 and 60 °C. The activity of colloid chitin degradation for Chi74 was the highest, followed by Chi66, Chi63 and Chi54. Ag+ reduced the activity of Chi74, Chi54, Chi63 and Chi66, but Mg2+ enhanced them. The effect of Ag+ and Mg2+ was more significant on the activity of Chi54 than on the activities of Chi63, Chi66 and Chi74.Conclusion
ChBDChi74 and FN3Chi74 domains play a role in exerting enzymatic activity and can improve the stability of chitinase.11.
Adones Sales Luana Ferreira Afonso Juliana Alves Americo Mauro de Freitas Rebelo Glaucia Maria Pastore Juliano Lemos Bicas 《Biotechnology letters》2018,40(3):561-567
Objective
To investigate the biocatalytic potential of Colletotrichum acutatum and Colletotrichum nymphaeae for monoterpene biotransformation.Results
C. acutatum and C. nymphaeae used limonene, α-pinene, β-pinene, farnesene, citronellol, linalool, geraniol, perillyl alcohol, and carveol as sole carbon and energy sources. Both species biotransformed limonene and linalool, accumulating limonene-1,2-diol and linalool oxides, respectively. α-Pinene was only biotransformed by C. nymphaeae producing campholenic aldehyde, pinanone and verbenone. The biotransformation of limonene by C. nymphaeae yielded 3.34–4.01 g limonene-1,2-diol l?1, depending on the substrate (R-(+)-limonene, S-(?)-limonene or citrus terpene (an agro-industrial by-product). This is among the highest concentrations already reported for this product.Conclusions
This is the first report on the biotransformation of these terpenes by Colletotrichum spp. and the biotransformation of limonene to limonene-1,2-diol possibly involves enzymes similar to those found in Grosmannia clavigera.12.
Changling Zhang Haifeng Pan Lingli Yao Wenna Bao Jinxin Wang Zhipeng Xie Jianguo Zhang 《Biotechnology letters》2016,38(8):1301-1306
Objectives
To enhance activity of cis-epoxysuccinate hydrolase from Klebsiella sp. BK-58 for converting cis-epoxysuccinate to tartrate.Results
By semi-saturation mutagenesis, all the mutants of the six important conserved residues almost completely lost activity. Then random mutation by error-prone PCR and high throughput screening were further performed to screen higher activity enzyme. We obtained a positive mutant F10D after screening 6000 mutations. Saturation mutagenesis on residues Phe10 showed that most of mutants exhibited higher activity than the wild-type, and the highest mutant was F10Q with activity of 812 U mg?1 (k cat /K m , 9.8 ± 0.1 mM?1 s?1), which was 230 % higher than that of wild-type enzyme 355 U mg?1 (k cat /K m , 5.3 ± 0.1 mM?1 s?1). However, the thermostability of the mutant F10Q slightly decreased.Conclusions
The catalytic activity of a cis-epoxysuccinate hydrolase was efficient improved by a single mutation F10Q and Phe10 might play an important role in the catalysis.13.
Camila Gazolla Volpiano Bruno Brito Lisboa Jackson Freitas Brilhante São José Andreia Mara Rotta de Oliveira Anelise Beneduzi Luciane Maria Pereira Passaglia Luciano Kayser Vargas 《Plant and Soil》2018,432(1-2):229-243
Aims
To identify Rhizobium strains’ ability to biocontrol Sclerotium rolfsii, a fungus that causes serious damage to the common bean and other important crops, 78 previously isolated rhizobia from common bean were assessed.Methods
Dual cultures, volatiles, indole-acetic acid (IAA), siderophore production and 16S rRNA sequencing were employed to select strains for pot and field experiments.Results
Thirty-three antagonistic strains were detected in dual cultures, 16 of which were able to inhibit ≥84% fungus mycelial growth. Antagonistic strains produced up to 36.5 μg mL?1 of IAA, and a direct correlation was verified between IAA production and mycelium inhibition. SEMIA 460 inhibited 45% of mycelial growth through volatile compounds. 16S rRNA sequences confirmed strains as Rhizobium species. In pot condition, common bean plants grown on S. rolfsii-infested soil and inoculated with SEMIA 4032, 4077, 4088, 4080, 4085, or 439 presented less or no disease symptoms. The most efficient strains under field conditions, SEMIA 439 and 4088, decreased disease incidence by 18.3 and 14.5% of the S. rolfsii-infested control.Conclusions
Rhizobium strains could be strong antagonists towards S. rolfsii growth. SEMIA 4032, 4077, 4088, 4080, 4085, and 439 are effective in the biological control of the collar rot of the common bean.14.
Andrelisse Arruda Viviane Castelo Branco Reis Vinícius Daniel Ferreira Batista Bruno Sahim Daher Luiza Cesca Piva Janice Lisboa De Marco Lidia Maria Pepe de Moraes Fernando Araripe Gonçalves Torres 《Biotechnology letters》2016,38(3):509-517
Objectives
To develop a new vector for constitutive expression in Pichia pastoris based on the endogenous glycolytic PGK1 promoter.Results
P. pastoris plasmids bearing at least 415 bp of PGK1 promoter sequences can be used to drive plasmid integration by addition at this locus without affecting cell growth. Based on this result, a new P. pastoris integrative vector, pPICK2, was constructed bearing some features that facilitate protein production in this yeast: a ~620 bp PGK1 promoter fragment with three options of restriction sites for plasmid linearization prior to yeast transformation: a codon-optimized α-factor secretion signal, a new polylinker, and the kan marker for vector propagation in bacteria and selection of yeast transformants.Conclusions
A new constitutive vector for P. pastoris represents an alternative platform for recombinant protein production and metabolic engineering purposes.15.
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.16.
Tatyana Laurinavichene Kestutis Laurinavichius Evgeny Shastik Anatoly Tsygankov 《Biotechnology letters》2018,40(2):309-314
Objectives
To prove the possibility of efficient starch photofermentation in co-culture of heterotrophic and phototrophic bacteria over prolonged period.Results
Repeated batch photofermentation of starch was demonstrated in co-culture Clostridium butyricum and Rhodobacter sphaeroides under microaerobic conditions. It continued 15 months without addition of new inoculum or pH regulation when using 4–5 g starch l?1 and 0.04 g yeast extract l?1. The complete degradation of starch without volatile fatty acids accumulation was shown in this co-culture. The average H2 yield of 5.2 mol/mol glucose was much higher than that in Clostridium monoculture. The species composition of co-culture was studied by q-PCR assay. The concentration of Clostridium cells in prolonged co-culture was lower than in monoculture and even in a single batch co-culture. This means that Clostridia growth was significantly limited whereas starch hydrolysis still took place.Conclusion
The prolonged repeated batch photofermentation of starch by co-culture C. butyricum and R. sphaeroides provided efficient H2 production without accumulation of organic acids under conditions of Clostridia limitation.17.
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 4.18.
Objectives
To investigate the outcomes of capsule lost on cell transformation efficiency and chemicals (1,3-propanediol, 2,3-butanediol, and 2-ketogluconic acid) production by Klebsiella pneumoniae.Results
The cps gene cluster showed low sequence homology with pathogenic strains. The wza is a highly conserved gene in the cps cluster that encodes an outer membrane protein. A non-capsulated mutant was constructed by deletion of wza. Phenotype studies demonstrated that non-capsulated cells were less buoyant and easy to sediment. The transformation efficiency of the non-capsulated mutant reached 6.4 × 105 CFU μg?1 DNA, which is 10 times higher than that of the wild strain. 52.2 g 1,3-propanediol L?1, 30.7 g 2,3-butanediol L?1, and 175.9 g 2-ketogluconic acid L?1 were produced by non-capsulated mutants, which were 10–40% lower compared to wild strain. Furthermore, viscosities of the three fermentation broths decreased to approximately 1.3 cP from the range of 1.8–2.2 cP.Conclusions
Non-capsulated K. pneumoniae mutants should allay concerns regarding biological safety, improve transformation efficiency, lower viscosity, and subsequently ameliorate the financial burden of the downstream process of chemicals production.19.
Aims
Despite extensive studies on effects of elevated CO2 concentration ([CO2]e) on plant growth, few studies have investigated the responses of native grassland plant species to [CO2]e in terms of nutrient acquisition.Methods
The effects of [CO2]e (769 ± 23 ppm) on Artemisia frigida and Stipa krylovii, two dominant species in Inner Mongolia steppe were investigated by growing them for 7 weeks in Open-Top Chambers (OTC).Results
Exposure to [CO2]e enhanced shoot and root growth of A. frigida and S. krylovii. Elevated [CO2] increased photosynthetic rates (Pn) by 34 % in A. frigida but decreased Pn by 52 % in S. krylovii. Moreover, root-secreted acid phosphatase activity in A. frigida was stimulated by [CO2]e, while exudation of malate from roots of S. krylovii was suppressed by [CO2]e. Exposure to [CO2]e led to a decrease in P concentration in shoots and roots of A. frigida and S. krylovii, but total amount of P accumulated in shoots and roots of both species was increased by [CO2]e.Conclusions
The two dominant species in temperate steppes differed in their responses to [CO2]e, such that A. frigida was more adapted to [CO2]e than S. krylovii under low availability of soil P.20.