Heterotrophic nitrification-aerobic denitrification performance of a novel strain, Pseudomonas sp. B-1, isolated from membrane aerated biofilm reactor.

A heterotrophic nitrification-aerobic denitrification (HN-AD) strain isolated from membrane aerated biofilm reactor (MABR) was identified as Pseudomonas sp. B-1, which could effectively utilize multiple nitrogen sources and preferentially consume NH4-N. The maximum degradation efficiencies of NO3-N, NO2-N and NH4-N were 98.04%, 94.84% and 95.74%, respectively. The optimal incubation time, shaking speed, carbon source, pH, temperature and C/N ratio were 60 h, 180 rpm, sodium succinate, 8, 30 °C and 25, respectively. The strain preferred salinity of 1.5% and resisted heavy metals in the order of Mn2+ > Co2+ > Zn2+ > Cu2+. It can be preliminarily speculated from the results of enzyme assay that the strain removed nitrogen via full nitrification-denitrification pathway. The addition of strain into the conventional MABR significantly intensified the HN-AD performance of the reactor. The relative abundance of the functional bacteria including Flavobacterium, Pseudomonas, Paracoccus, Azoarcus and Thauera was obviously increased after the bioaugmentation. Besides, the expression of the HN-AD related genes in the biofilm was also strengthened. Thus, strain B-1 had great application potential in nitrogen removal process.

Enhancement of the polyethylene terephthalate and mono-(2-hydroxyethyl) terephthalate degradation activity of Ideonella sakaiensis PETase by an electrostatic interaction-based strategy.

The serious environmental pollution that came up with the continuously growing demand for polyethylene terephthalate (PET) has attracted global concern. The IsPETase which has shown the highest PET degradation activity under ambient temperature is a promising enzyme for PET biodegradation, while poor thermostability limited its practical application. Herein, an electrostatic interaction-based strategy was applied for rational design of IsPETase towards enhanced thermostability. The IsPETaseI139R variant displayed the highest Tm value of 56.4 °C and 3.6-times higher PET degradation activity. Molecular simulations demonstrated that the introduction of salt bridges stabilized the local structures, resulting in robust thermostability. Meanwhile, the IsPETaseS92K/D157E/R251A not only exhibited higher thermostability but also showed a 1.74-fold kcat increase towards mono-(2-hydroxyethyl) terephthalate, which ultimately achieved PET depolymerization to complete monomer TPA. Collectively, the electrostatic interaction-based strategy, together with the derived IsPETase variants, could help promote the bio-recycle of PET, reducing the severe global burden of PET waste.

Utilization of whey powder as substrate for low-cost preparation of ß-galactosidase as main product, and ethanol as by-product, by a litre-scale integrated process.

Whey powder, a by-product of dairy industry, is an attractive raw material for value-added products. In this study, utilization of whey powder as substrate for low-cost preparation of ß-galactosidase as main product and ethanol as by-product were investigated by a litre-scale integrated strategy, encompassing fermentation, isolation, permeabilization and spray drying. Firstly, through development of low-cost industrial culture and fed-batch strategies by Kluyveromyces lactis, 119.30U/mL ß-galactosidase activity and 16.96mg/mL by-product ethanol were achieved. Afterward, an up-dated mathematic model for the recycling permeabilization was established successfully and 30.4g cells sediment isolated from 5L fermentation broth were permeabilized completely by distilled ethanol from broth supernatant. Then ß-galactosidase product with 5.15U/mg from protection of gum acacia by spray drying was obtained. Furthermore, by-product ethanol with 31.08% (v/v) was achieved after permeabilization. Therefore, the integrated strategy using whey powder as substrate is a feasible candidate for industrial-scale implementation.

Utilization of biodiesel by-product as substrate for high-production of ß-farnesene via relatively balanced mevalonate pathway in Escherichia coli.

Farnesene has been identified as suitable jet fuel substitutes and metabolic engineering for microbial production of farnesene is an alternative and attractive route. In this study, due to accumulation of toxic intermediate isopentenyl pyrophosphate (IPP), an engineered Escherichia coli strain harboring heterologous mevalonate pathway produced only 4.11mg/L ß-farnesene. Through higher-level expression of isopentenyl diphosphate isomerase and farnesyl diphosphate synthase to minimize the accumulated IPP, another engineered strain with relatively balanced mevalonate pathway was constructed and had the highest production of ß-farnesene to date (8.74g/L) by Escherichia coli in a lab bioreactor. Furthermore, this is the first report on utilization of biodiesel by-product (simple purification) as substrate for high-production of ß-farnesene by the engineered strain optimized and ß-farnesene concentration reached 2.83g/L in a lab bioreactor. Therefore, the engineered strain optimized could be used as a platform host for high-production of other terpenoids using biodiesel by-product as substrate.

Development of a novel integrated process for co-production of ß-galactosidase and ethanol using lactose as substrate.

A novel integrated process was developed successfully for co-production of ß-galactosidase and ethanol using lactose as substrate, containing fermentation (Kluyveromyces lactis), isolation, permeabilization (a new recycling process) and spray drying. Firstly, a new fed-batch strategy optimized co-produced ß-galactosidase at 105.91U/mL and ethanol at 32.16mg/mL, 4.40-fold and 10.82-fold increase over the results from initial conditions, respectively. Then a new mathematic model for the recycling permeabilization was established successfully. As expected, the total cells sediment from isolation of the fed-batch culture was permeabilized completely by distilled ethanol from broth supernatant. More amazedly, the specific activity of ß-galactosidase product by spray drying the permeabilized cells reached 2.61U/mg, meeting the demand of commercial products. Furthermore, the ethanol product at 33.8% (v/v) was obtained from the novel integrated process, which could be applied for various applications. To conclude, the novel integrated process might be a feasible strategy to scale up for industrialization.