Yongze Wang

Homofermentative production of optically pure L-lactic acid from xylose by genetically engineered Escherichia coli B

BackgroundPolylactic acid (PLA), a biodegradable polymer, has the potential to replace (at least partially) traditional petroleum-based plastics, minimizing “white pollution”. However, cost-effective production of optically pure L-lactic acid is needed to achieve the full potential of PLA. Currently, starch-based glucose is used for L-lactic acid fermentation by lactic acid bacteria. Due to its competition with food resources, an alternative non-food substrate such as cellulosic biomass is needed for L-lactic acid fermentation. Nevertheless, the substrate (sugar stream) derived from cellulosic biomass contains significant amounts of xylose, which is unfermentable by most lactic acid bacteria. However, the microorganisms that do ferment xylose usually carry out heterolactic acid fermentation. As a result, an alternative strain should be developed for homofermentative production of optically pure L-lactic acid using cellulosic biomass.ResultsIn this study, an ethanologenic Escherichia coli strain, SZ470 (ΔfrdBC ΔldhA ΔackA ΔpflB ΔpdhR ::pflBp6-acEF-lpd ΔmgsA), was reengineered for homofermentative production of L-lactic acid from xylose (1.2 mole xylose = > 2 mole L-lactic acid), by deleting the alcohol dehydrogenase gene (adhE) and integrating the L-lactate dehydrogenase gene (ldhL) of Pediococcus acidilactici. The resulting strain, WL203, was metabolically evolved further through serial transfers in screw-cap tubes containing xylose, resulting in the strain WL204 with improved anaerobic cell growth. When tested in 70 g L-1 xylose fermentation (complex medium), WL204 produced 62 g L-1 L-lactic acid, with a maximum production rate of 1.631 g L-1 h-1 and a yield of 97% based on xylose metabolized. HPLC analysis using a chiral column showed that an L-lactic acid optical purity of 99.5% was achieved by WL204.ConclusionsThese results demonstrated that WL204 has the potential for homofermentative production of L-lactic acid using cellulosic biomass derived substrates, which contain a significant amount of xylose.

Engineering and adaptive evolution of Escherichia coli W for L-lactic acid fermentation from molasses and corn steep liquor without additional nutrients.

The D-lactic acid producing strain, Escherichia coli HBUT-D, was reengineered for L(+)-lactic acid fermentation by replacing the D-lactate dehydrogenase gene (ldhA) with an L(+)-lactate dehydrogenase gene (ldhL) from Pedicoccus acidilactici, followed by adaptive evolution in sucrose. The resulting strain, WYZ-L, has enhanced expression of the sucrose operon (cscA and cscKB). In 100 g L(-1) of sucrose fermentation using mineral salt medium, WYZ-L produced 97 g L(-1) of l(+)-lactic acid, with a yield of 90%, a maximum productivity of 3.17 g L(-1)h(-1) and an optical purity of greater than 99%. In fermentations using sugarcane molasses and corn steep liquor without additional nutrients, WYZ-L produced 75 g L(-1) of l(+)-lactic acid, with a yield of 85%, a maximum productivity of 1.18 g L(-1)h(-1), and greater than 99% optical purity. These results demonstrated that WYZ-L has the potential to use waste molasses and corn steep liquor as a resource for L(+)-lactic acid fermentation.

Fermentation characteristics of engineered Escherichia coli for succinic acid production

Succinic acid produced by microbial fermentation has much environmental benefits and attracted great interest currently. Among them, Escherichia coli have been chosen as good succinic acid producing strains because of their advantages. Normally, no more than 0.2 mol of succinate is formed per mol of glucose consumed by E. coli. In order to improve the yield of succinic acid, a genetically modified E. coli JH204 was constructed through RED recombination system and the flipase recognition target (FRT) site-specific recombinant technology, and multi-genes were knocked out from wild strain E. coli W, including ldhA, adhE, pflB, poxB and ackA to block the metabolic competition pathways. In this paper, the fermentation characteristics of the succinic acid production by engineered strain was investigated. The mutant carries out a favorable fermentation of glucose in 10L fermenter, and the production of succinic acid was approximately 40.08g/L, and the productivity and yield were 0.83gL-1h-1 and 0.72g/g glucose respectively. Acetic acid and lactic acid were the only products detected and accumulated to concentrations of 4.53g/L and 0.08g/L, respectively. And formic acid and ethanol were not detected in the medium of the fermentation.

Activated alumina enhanced biogas production from corn straw hydrolysate

The effects of the activated alumina on biogas production from corn straw hydrolysate were investigated in an up-flow anaerobic sludge blanket (UASB). The biogas fermentation was initiated by inoculation of the mixture of activated alumina and sludge (1 to 19 ratios) into the UASB, and operated at 37 _ for two month. The results showed that the activated alumina enabled 10 days earlier (10 days vs 20 days) of the start-up period for maximum methane production and increased the methane content of the biogas produced by 23% (86% vs 70%) when compared to those of the control With the added activated alumina, the biogas fermentation of corn straw hydrolysate achieved chemical oxygen demand (COD) removal rate, organic load rate (OLR), methane yield, and the percentage of methane in the biogas produced by 82.3%, 15.01 g/L d COD, 304 ml CH4/g COD, and 86%, respectively.