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在大肠杆菌中展示巨大芽孢杆菌的α-淀粉酶,用于将淀粉生物转化为氢气、乙醇和琥珀酸。

Autodisplay of alpha amylase from Bacillus megaterium in E. coli for the bioconversion of starch into hydrogen, ethanol and succinic acid.

机构信息

División de Biología Molecular, Instituto Potosino de Investigación Científica y Tecnológica, A.C. Camino a la Presa San José 2055, Col. Lomas 4ª Sección, C.P. 78216, San Luis Potosí, México.

División de Biología Molecular, Instituto Potosino de Investigación Científica y Tecnológica, A.C. Camino a la Presa San José 2055, Col. Lomas 4ª Sección, C.P. 78216, San Luis Potosí, México.

出版信息

Enzyme Microb Technol. 2020 Mar;134:109477. doi: 10.1016/j.enzmictec.2019.109477. Epub 2019 Nov 19.


DOI:10.1016/j.enzmictec.2019.109477
PMID:32044024
Abstract

In this work, the expression of an α-amylase from Bacillus megaterium on the cell surface of Escherichia coli strains WDHA (Δ hycA and Δ ldhA) and WDHFP (Δ hycA, Δ frdD and Δ pta) by the autodisplay adhesin involved in diffuse adherence (AIDA) system was carried out with the purpose to confer the ability to E. coli strains to degrade starch and thus produce hydrogen, ethanol and succinic acid. For the characterization of the biocatalyst, the effect of temperature (30-70 °C), pH (3-6) and CaCl concentration (0-25 mM), as well as the thermostability of the biocatalyst (55-80 °C) at several time intervals (15-60 min) were evaluated. The results showed that the biocatalyst had a maximum activity at 55 °C and pH 4.5. Calcium was required for the activity as well for the thermal stability of the biocatalyst. The calculated Vmax and Km values were 0.24 U/cm and 5.8 mg/cm, respectively. Furthermore, a set of anaerobic batch fermentations was carried out using 10 g/dm of starch and 1 g/dm of glucose as carbon sources in 120 cm serological bottles, using WDHA and WDHFP strains harboring the pAIDA-amyA plasmid. The hydrogen production for WDHA was 1056.06 cm/dm and the succinic acid yield was 0.68 g/g, whereas WDHFP strain produced 1689.68 cm/dm of hydrogen and an ethanol yield of 0.28 g/g. This work represents a promising strategy to improve the exploitation of starchy biomass for the production of biofuels (hydrogen and ethanol) or succinate without the need of a pre-saccharification process.

摘要

在这项工作中,通过自展示黏附素(diffuse adherence (AIDA) 系统)将巨大芽孢杆菌的α-淀粉酶表达在大肠杆菌菌株 WDHA(ΔhycA 和 ΔldhA)和 WDHFP(ΔhycA、ΔfrdD 和 Δpta)的细胞表面上,目的是赋予大肠杆菌菌株降解淀粉的能力,从而生产氢气、乙醇和琥珀酸。为了对生物催化剂进行表征,评估了温度(30-70°C)、pH(3-6)和 CaCl2浓度(0-25 mM)以及生物催化剂的热稳定性(55-80°C)在几个时间间隔(15-60 分钟)的影响。结果表明,生物催化剂在 55°C 和 pH 4.5 时具有最大活性。钙对活性和生物催化剂的热稳定性都是必需的。计算出的 Vmax 和 Km 值分别为 0.24 U/cm 和 5.8 mg/cm。此外,在 120 cm 血清瓶中使用 10 g/dm 的淀粉和 1 g/dm 的葡萄糖作为碳源进行了一组厌氧分批发酵,使用携带 pAIDA-amyA 质粒的 WDHA 和 WDHFP 菌株。WDHA 的氢气产量为 1056.06 cm/dm,琥珀酸产率为 0.68 g/g,而 WDHFP 菌株产生了 1689.68 cm/dm 的氢气和 0.28 g/g 的乙醇。这项工作代表了一种有前途的策略,可以在无需进行预糖化过程的情况下,提高淀粉生物质生产生物燃料(氢气和乙醇)或琥珀酸的利用效率。

相似文献

[1]
Autodisplay of alpha amylase from Bacillus megaterium in E. coli for the bioconversion of starch into hydrogen, ethanol and succinic acid.

Enzyme Microb Technol. 2019-11-19

[2]
Probing the function of C-terminal region of recombinant α-amylase BmaN1 from NL3.

Microbiol Spectr. 2024-10-3

[3]
Production of itaconic acid in Escherichia coli expressing recombinant α-amylase using starch as substrate.

J Biosci Bioeng. 2015-5

[4]
Direct fermentation of raw starch using a Kluyveromyces marxianus strain that expresses glucoamylase and alpha-amylase to produce ethanol.

Biotechnol Prog. 2014

[5]
Direct bioethanol production by amylolytic yeast Candida albicans.

Lett Appl Microbiol. 2015-3

[6]
Production of ethanol from starch by free and immobilized Candida tropicalis in the presence of alpha-amylase.

Bioresour Technol. 2007-10

[7]
Fermentation of starch by Klebsiella oxytoca p2, containing plasmids with alpha-amylase and pullulanase genes.

Biotechnol Bioeng. 1999-12-20

[8]
A chimeric α-amylase engineered from Bacillus acidicola and Geobacillus thermoleovorans with improved thermostability and catalytic efficiency.

J Ind Microbiol Biotechnol. 2016-4

[9]
Efficient carbon dioxide utilization and simultaneous hydrogen enrichment from off-gas of acetone-butanol-ethanol fermentation by succinic acid producing Escherichia coli.

Bioresour Technol. 2016-4-19

[10]
Fermentation optimization and amylase activity of endophytic Bacillus velezensis D1 isolated from corn seeds.

J Appl Microbiol. 2022-5

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[2]
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[3]
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[4]
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