Stable, efficient, and cost-effective system for the biosynthesis of recombinant bacterial cellulose in Escherichia coli DH5α platform.

BACKGROUND: Owing to its remarkable mechanical properties that surpass the plant-based cellulose, bacterial cellulose production has been targeted for commercialization during the last few years. However, the large-scale production of cellulose is generally limited by the slow growth of producing strains and low productivity which ultimately makes the commercial production of cellulose using the conventional strains non cost-effective. In this study, we developed a novel plasmid-based expression system for the biosynthesis of cellulose in E. coli DH5α and assessed the cellulose productivity relative to the typically used E. coli BL21 (DE) expression strain. RESULTS: No production was detected in BL21 (DE3) cultures upon expression induction; however, cellulose was detected in E. coli DH5α as early as 1 h post-induction. The total yield in induced DH5α cultures was estimated as 200 ± 5.42 mg/L (dry weight) after 18 h induction, which surpassed the yield reported in previous studies and even the wild-type Gluconacetobacter xylinum BRC5 under the same conditions. As confirmed with electron microscope micrograph, E. coli DH5α produced dense cellulose fibers with ~ 10 µm diameter and 1000-3000 µm length, which were remarkably larger and more crystalline than that typically produced by G. hansenii. CONCLUSIONS: This is the first report on the successful cellulose production in E. coli DH5α which is typically used for plasmid multiplication rather than protein expression, without the need to co-express cmcax and ccpAx regulator genes present in the wild-type genome upstream the bcs-operon, and reportedly essential for the biosynthesis.

Statistical optimization and characterization of a biocellulose produced by local Egyptian isolate Komagataeibacter hansenii AS.5.

Optimization of the culture parameters used for biocellulose (BC) production by a previously isolated bacterial strain (Komagataeibacter hansenii AS.5) was carried out. The effect of nine culture parameters on BC production was evaluated by implementing the Plackett-Burman design, and the results revealed that, the most significant variables affecting BC production were MgSO4, ethanol, pH and yeast extract. A three-level and four-factor Box-Behnken design was applied to determine the optimum level of each significant variable. According to the results of the Plackett-Burman (PBD) and Box-Behnken designs (BBD), the following medium composition and parameters were calculated to be optimum (g/l): glucose 25, yeast extract 13, MgSO4 0.15, KH2PO4 2, ethanol 7.18 ml/l, pH 5.5, inoclume size 7%, cultivation temperature 20 °C and incubation time 9 days. Characterization of purified BC was performed to determine the network morphology by scanning electron microscopy, crystallinity by X-ray diffraction, chemical structure and functional groups by Fourier-transform infrared spectroscopy, thermal stability by thermogravimetric analysis and mechanical properties such as Young's modulus, tensile strength and elongation at beak % of BC.