Dataset Information

Light induced expression of ?-glucosidase in Escherichia coli with autolysis of cell.

ABSTRACT: BACKGROUND:?-Glucosidase has attracted substantial attention in the scientific community because of its pivotal role in cellulose degradation, glycoside transformation and many other industrial processes. However, the tedious and costly expression and purification procedures have severely thwarted the industrial applications of ?-glucosidase. Thus development of new strategies to express ?-glucosidases with cost-effective and simple procedure to meet the increasing demands on enzymes for biocatalysis is of paramount importance. RESULTS:Light activated cassette YF1/FixJ and the SRRz lysis system were successfully constructed to produce Bgl1A(A24S/F297Y), a mutant ?-glucosidase tolerant to both glucose and ethanol. By optimizing the parameters for light induction, Bgl1A(A24S/F297Y) activity reached 33.22 ± 2.0 U/mL and 249.92 ± 12.25 U/mL in 250-mL flask and 3-L fermentation tank, respectively, comparable to the controls of 34.02 ± 1.96 U/mL and 322.21 ± 10.16 U/mL under similar culture conditions with IPTG induction. To further simplify the production of our target protein, the SRRz lysis gene cassette from bacteriophage Lambda was introduced to trigger cell autolysis. As high as 84.53 ± 6.79% and 77.21 ± 4.79% of the total ?-glucosidase were released into the lysate after cell autolysis in 250 mL flasks and 3-L scale fermentation with lactose as inducer of SRRz. In order to reduce the cost of protein purification, a cellulose-binding module (CBM) from Clostridium thermocellum was fused into the C-terminal of Bgl1A(A24S/F297Y) and cellulose was used as an economic material to adsorb the fusion enzyme from the lysate. The yield of the fusion protein could reach 92.20 ± 2.27% after one-hour adsorption at 25 °C. CONCLUSIONS:We have developed an efficient and inexpensive way to produce ?-glucosidase for potential industrial applications by using the combination of light induction, cell autolysis, and CBM purification strategy.

SUBMITTER: Chang F

PROVIDER: S-EPMC5688802 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Light induced expression of β-glucosidase in Escherichia coli with autolysis of cell.

Chang Fei F Zhang Xianbing X Pan Yu Y Lu Youxue Y Fang Wei W Fang Zemin Z Xiao Yazhong Y

BMC biotechnology 20171107 1

<h4>Background</h4>β-Glucosidase has attracted substantial attention in the scientific community because of its pivotal role in cellulose degradation, glycoside transformation and many other industrial processes. However, the tedious and costly expression and purification procedures have severely thwarted the industrial applications of β-glucosidase. Thus development of new strategies to express β-glucosidases with cost-effective and simple procedure to meet the increasing demands on enzymes for ...[more]

PMID: 29115967

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Project description:BACKGROUND: Solar energy is the ultimate energy source on the Earth. The conversion of solar energy into fuels and energy sources can be an ideal solution to address energy problems. The recent discovery of proteorhodopsin in uncultured marine ?-proteobacteria has made it possible to construct recombinant Escherichia coli with the function of light-driven proton pumps. Protons that translocate across membranes by proteorhodopsin generate a proton motive force for ATP synthesis by ATPase. Excess protons can also be substrates for hydrogen (H(2)) production by hydrogenase in the periplasmic space. In the present work, we investigated the effect of the co-expression of proteorhodopsin and hydrogenase on H(2) production yield under light conditions. RESULTS: Recombinant E. coli BL21(DE3) co-expressing proteorhodopsin and [NiFe]-hydrogenase from Hydrogenovibrio marinus produced ~1.3-fold more H(2) in the presence of exogenous retinal than in the absence of retinal under light conditions (70 ?mole photon/(m2·s)). We also observed the synergistic effect of proteorhodopsin with endogenous retinal on H(2) production (~1.3-fold more) with a dual plasmid system compared to the strain with a single plasmid for the sole expression of hydrogenase. The increase of light intensity from 70 to 130 ?mole photon/(m(2)·s) led to an increase (~1.8-fold) in H(2) production from 287.3 to 525.7 mL H(2)/L-culture in the culture of recombinant E. coli co-expressing hydrogenase and proteorhodopsin in conjunction with endogenous retinal. The conversion efficiency of light energy to H(2) achieved in this study was ~3.4%. CONCLUSION: Here, we report for the first time the potential application of proteorhodopsin for the production of biohydrogen, a promising alternative fuel. We showed that H(2) production was enhanced by the co-expression of proteorhodopsin and [NiFe]-hydrogenase in recombinant E. coli BL21(DE3) in a light intensity-dependent manner. These results demonstrate that E. coli can be applied as light-powered cell factories for biohydrogen production by introducing proteorhodopsin.

Dataset Information

Light induced expression of ?-glucosidase in Escherichia coli with autolysis of cell.

Publications

Light induced expression of β-glucosidase in Escherichia coli with autolysis of cell.

Similar Datasets

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