Three Novel <i>Escherichia coli</i> Vectors for Convenient and Efficient Molecular Biological Manipulations

Gao, Herui; Qi, Xianghui; Hart, Darren J.; Gao, Song; Wang, Hongling; Xu, Shuangyan; Zhang, Yifeng; Liu, Xia; Liu, Yifei; An, Yi

doi:10.1021/acs.jafc.8b01960

Cited by 17 publications

(11 citation statements)

References 41 publications

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“…They were routinely cultured in Luria-Bertani (LB) medium containing Kanamycin (50 µg/mL) in a rotary shaker at 37 • C and 200 rpm. The plasmid pANY1 was obtained from the Shenyang Agricultural University, China (Gao et al, 2018;Liu et al, 2018), and was used as the expression vector.…”

Section: Bacterial Strains and Materialsmentioning

confidence: 99%

Biocatalytic Synthesis of D-Allulose Using Novel D-Tagatose 3-Epimerase From Christensenella minuta

et al. 2020

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D-allulose, which is one of the important rare sugars, has gained significant attention in the food and pharmaceutical industries as a potential alternative to sucrose and fructose. Enzymes belonging to the D-tagatose 3-epimerase (DTEase) family can reversibly catalyze the epimerization of D-fructose at the C3 position and convert it into D-allulose by a good number of naturally occurring microorganisms. However, microbial synthesis of D-allulose is still at its immature stage in the industrial arena, mostly due to the preference of slightly acidic conditions for Izumoring reactions. Discovery of novel DTEase that works at acidic conditions is highly preferred for industrial applications. In this study, a novel DTEase, DTE-CM, capable of catalyzing D-fructose into D-allulose was applications. In this study, a novel DTEase, DTE-CM, capable of catalyzing D-fructose into D-allulose was DTE-CM on D-fructose was found to be remarkably influenced and modulated by the type of metal ions (co-factors). The DTE-CM on D-fructose was found to be remarkably influenced and modulated by the type of metal ions (co-factors). The 50°C from 0.5 to 3.5 h at a concentration of 0.1 mM. The enzyme exhibited its maximum catalytic activity on D-fructose at pH 6.0 and 50°C from 0.5 to 3.5 h at a concentration of 0.1 mM. The enzyme exhibited its maximum catalytic activity on -fructose at pH 6.0 and 50°C with a Kcat/Km value of 45 mM−1min−1. The 500 g/L D-fructose, which corresponded to 30% conversion rate. With these interesting catalytic properties, this enzyme could be a promising candidate for industrial biocatalytic applications.

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Section: Bacterial Strains and Materialsmentioning

confidence: 99%

Biocatalytic Synthesis of D-Allulose Using Novel D-Tagatose 3-Epimerase From Christensenella minuta

et al. 2020

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“…Then, the purified protein was eluted after incubating the column with 200 μL of pre-cooling cleaving buffer (phosphate-buffered saline buffer supplemented with 40 mmol/L bis-Tris, pH 7.0) for 12 h at 37 °C. As a comparison, the pfLamA protein was purified with the traditional ELP-intein-based method . The protein samples were checked by SDS-PAGE, and activities of pfLamA samples were determined by DNS assay as described above.…”

Section: Methodsmentioning

confidence: 99%

“…The activity of pfLamA originating from pANY4-T7-pfLamA was defined as 100%, and the ratio between the activity of each sample and the activity of pfLamA originating from pANY4-T7-pfLamA was shown as relative activity (%). The pfLamA gene of the bestperforming pANY4-pL18-pfLamA and pANY4-pR18/pL18-pfLamA plasmids was replaced using the ccdB cassette of the pANY2 plasmid 26 to construct the plasmids pANY4-pL18-ccdB and pANY4-pR18/pL18-ccdB, respectively. The plasmids pANY4-pL18-ccdB and pANY4-pR18/pL18-ccdB have been deposited to Addgene (nos.…”

Section: Methodsmentioning

confidence: 99%

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Efficient Molecular Biological Manipulations with Improved Strategies Based on Novel Escherichia coli Vectors

Zhang

Gao

et al. 2021

J. Agric. Food Chem.

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In this study, some novel plasmids have been constructed for flexible and zero-background molecular cloning, more efficient expression, and purification of proteins with improved strategies. The plasmids pANY4-pL18-ccdB and pANY4-pR18/ pL18-ccdB have different promoters in the complementary DNA strands. Therefore, recombinant plasmids for either isopropyl-β-Dthiogalactoside-induced or temperature-induced protein expression could be simultaneously constructed in a single molecular cloning process for parallel comparison. Intriguingly, the mutated pL18 and pR18/pL18 promoters performed similar to or even better than the T7 promoter when used for promoting the expression of the GFP or pfLamA enzyme. Moreover, the plasmid pANY8 containing the His-elastin-like polypeptide (ELP)-intein multifunctional tag was constructed, and special purification protocol was designed to obtain purified proteins without the requirement of time-consuming dialysis steps to remove imidazole and high concentration of salt ions. Additionally, the urea-based denaturation and refolding processes can be conveniently integrated into the ELP-mediated precipitation protocol for purification of insoluble inclusion bodies, omitting the time-consuming dialysis steps.

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“…It was cultivated anaerobically in TPY fluid Medium at 37 • C without agitation. pANY1 plasmid was obtained from Shenyang Agricultural University, China (Gao et al, 2018;Liu et al, 2018), which was used as the expression vector. E. coli BL21(DE3) was obtained from the Stratagene (California, United States).…”

Section: Bacterial Strains and Materialsmentioning

confidence: 99%

Exploring a Highly D-Galactose Specific L-Arabinose Isomerase From Bifidobacterium adolescentis for D-Tagatose Production

Zhang

Parvez

et al. 2020

Front. Bioeng. Biotechnol.

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D-Galactose-specific L-arabinose isomerase (L-AI) would have much potential for the enzymatic conversion of D-Galactose into D-tagatose, while most of the reported L-AIs are L-arabinose specific. This study explored a highly D-Galactose-specific L-AI from Bifidobacterium adolescentis (BAAI) for the production of D-tagatose. In the comparative protein-substrate docking for D-Galactose and L-arabinose, BAAI showed higher numbers of hydrogen bonds in D-Galactose-BAAI bonding site than those found in L-arabinose-BAAI bonding site. The activity of BAAI was 24.47 U/mg, and it showed good stability at temperatures up to 65 • C and a pH range 6.0-7.5. The K m , V max , and K cat /K m of BAAI were found to be 22.4 mM, 489 U/mg and 9.3 mM −1 min −1 , respectively for D-Galactose, while the respective values for L-arabinose were 40.2 mM, 275.1 U/mg, and 8.6 mM −1 min −1 . Enzymatic conversion of D-Galactose into D-tagatose by BAAI showed 56.7% conversion efficiency at 55 • C and pH 6.5 after 10 h.

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Three Novel Escherichia coli Vectors for Convenient and Efficient Molecular Biological Manipulations

Cited by 17 publications

References 41 publications

Biocatalytic Synthesis of D-Allulose Using Novel D-Tagatose 3-Epimerase From Christensenella minuta

Biocatalytic Synthesis of D-Allulose Using Novel D-Tagatose 3-Epimerase From Christensenella minuta

Efficient Molecular Biological Manipulations with Improved Strategies Based on Novel Escherichia coli Vectors

Exploring a Highly D-Galactose Specific L-Arabinose Isomerase From Bifidobacterium adolescentis for D-Tagatose Production

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