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Autodisplay of alpha amylase from Bacillus megaterium in E. coli for the bioconversion of starch into hydrogen, ethanol and succinic acid
Enzyme and Microbial Technology ( IF 3.4 ) Pub Date : 2020-03-01 , DOI: 10.1016/j.enzmictec.2019.109477
Ana K Gutiérrez-García 1 , Cecilia Lizeth Alvarez-Guzmán 1 , Antonio De Leon-Rodriguez 1
Affiliation  

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 CaCl2 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/cm3 and 5.8 mg/cm3, respectively. Furthermore, a set of anaerobic batch fermentations was carried out using 10 g/dm3 of starch and 1 g/dm3 of glucose as carbon sources in 120 cm3 serological bottles, using WDHA and WDHFP strains harboring the pAIDA-amyA plasmid. The hydrogen production for WDHA was 1056.06 cm3/dm3 and the succinic acid yield was 0.68 g/gstarch, whereas WDHFP strain produced 1689.68 cm3/dm3 of hydrogen and an ethanol yield of 0.28 g/gstarch. 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.

中文翻译:

来自大肠杆菌中巨大芽孢杆菌的 α 淀粉酶的自动展示,用于将淀粉生物转化为氢、乙醇和琥珀酸

在这项工作中,来自巨大芽孢杆菌的 α-淀粉酶在大肠杆菌菌株 WDHA(Δ hycA 和 Δ ldhA)和 WDHFP(Δ hycA、Δ frdD 和 Δ pta)细胞表面通过参与弥散粘附的自展示粘附素表达(AIDA) 系统的目的是赋予大肠杆菌菌株降解淀粉的能力,从而产生氢气、乙醇和琥珀酸。对于生物催化剂的表征,温度 (30-70 °C)、pH (3-6) 和 CaCl2 浓度 (0-25 mM) 的影响,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)进行评估。结果表明,该生物催化剂在 55 °C 和 pH 4.5 时具有最大活性。生物催化剂的活性以及热稳定性都需要钙。计算出的 Vmax 和 Km 值为 0。分别为 24 U/cm3 和 5.8 mg/cm3。此外,使用含有 pAIDA-amyA 质粒的 WDHA 和 WDHFP 菌株,在 120 cm3 血清学瓶中使用 10 g/dm3 淀粉和 1 g/dm3 葡萄糖作为碳源进行了一组厌氧分批发酵。WDHA 的产氢量为 1056.06 cm3/dm3,琥珀酸产率为 0.68 g/g 淀粉,而 WDHFP 菌株产氢量为 1689.68 cm3/dm3,乙醇产率为 0.28 g/g 淀粉。这项工作代表了一种有前景的策略,可以在不需要预糖化过程的情况下,改进淀粉类生物质的开发,用于生产生物燃料(氢和乙醇)或琥珀酸盐。使用含有 pAIDA-amyA 质粒的 WDHA 和 WDHFP 菌株,在 120 cm3 血清瓶中使用 10 g/dm3 淀粉和 1 g/dm3 葡萄糖作为碳源进行一组厌氧分批发酵。WDHA 的产氢量为 1056.06 cm3/dm3,琥珀酸产率为 0.68 g/g 淀粉,而 WDHFP 菌株产氢量为 1689.68 cm3/dm3,乙醇产率为 0.28 g/g 淀粉。这项工作代表了一种有前景的策略,可以在不需要预糖化过程的情况下,改进淀粉类生物质的开发,用于生产生物燃料(氢和乙醇)或琥珀酸盐。使用含有 pAIDA-amyA 质粒的 WDHA 和 WDHFP 菌株,在 120 cm3 血清瓶中使用 10 g/dm3 淀粉和 1 g/dm3 葡萄糖作为碳源进行一组厌氧分批发酵。WDHA 的产氢量为 1056.06 cm3/dm3,琥珀酸产率为 0.68 g/g 淀粉,而 WDHFP 菌株产氢量为 1689.68 cm3/dm3,乙醇产率为 0.28 g/g 淀粉。这项工作代表了一种有前景的策略,可以在不需要预糖化过程的情况下,改进淀粉类生物质的开发,用于生产生物燃料(氢和乙醇)或琥珀酸盐。68 cm3/dm3的氢气和0.28g/g淀粉的乙醇产率。这项工作代表了一种有前景的策略,可以在不需要预糖化过程的情况下,改进淀粉类生物质的开发,用于生产生物燃料(氢和乙醇)或琥珀酸盐。68 cm3/dm3的氢气和0.28g/g淀粉的乙醇产率。这项工作代表了一种有前景的策略,可以在不需要预糖化过程的情况下,改进淀粉类生物质的开发,用于生产生物燃料(氢和乙醇)或琥珀酸盐。
更新日期:2020-03-01
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