Metabolic engineering of an E. coli ndh knockout strain for PHB production from mixed glucose–xylose feedstock

Abstract

BACKGROUND: Poly(3-hydroxybutyrate) (PHB), which is completely biodegradable, is considered a potential candidate to replace a number of petroleum-derived polymers due to similar mechanical properties. In a previous study, inactivation of ndh gene in E. coli, which encodes the NDH-II dehydrogenase, resulted in significantly increased PHB production from either glucose or xylose as substrate. RESULTS:In this study, the xylose isomerase (EC:5.3.1.5), xylulokinase (EC:2.7.1.17) and the arabinose/xylose transport protein from Bacillus subtilis 168 (encoded by xylA, xylB and araE, respectively) were co-expressed in the ndh knockout strain, E. coli LJ03(pBHR68), which harbors the PHB biosynthesis genes from Ralstonia eutropha. The resulting strain E. coli LJ03(pBHR68+pM-ABE) was able to simultaneously utilize glucose and xylose to accumulate PHB. In flask cultivation, 3.67 g L −1 PHB was produced from a glucose-xylose mixture (10 g L −1 glucose and 5 g L −1 xylose), which was 2.09-fold higher than the production of the control strain E. coli JM109(pBHR68+pM-ABE). Ultimately, PHB production in fed-batch fermentation reached a maximum titer of 21.0 g L −1 , representing a 1.93-fold increase relative to the control strain. CONCLUSION: Results indicated that the engineered E. coli LJ03 strain is significantly more efficient than the parent strain E. coli JM109 in producing PHB from mixed glucose-xylose feedstock. To the best of our knowledge, this is the first study describing the implementation of an exogenous xylose utilization pathway in E. coli for the production of PHB from mixed glucose-xylose feedstock -a model of lignocellulosic hydrolysate.

Construction of Efficient Platform Escherichia coli Strains for Polyhydroxyalkanoate Production by Engineering Branched Pathway

“…To date, PHA has been shown to be produced by many microorganisms, including R. eutropha , A. latus , and recombinant E. coli using diverse inexpensive carbon sources [4,7,8]. In addition to pathway construction, many studies also focus on enhancing the supply of acetyl-CoA and/or NADPH [10].…”

“…PHA production in recombinant E. coli is not only a matter of pathway construction but is also affected by many other factors, such as acetyl-CoA and NADPH. Several strategies have focused on enhancing the supply of NADPH and/or acetyl-CoA to improve PHA production [10]. First, acetyl-CoA is a crucial intermediate for PHA production and can directly increase 3-hydroxybuturyl-CoA levels and cell growth.…”

Escherichia coli as a host for metabolic engineering

“…After facile hydrolysis of hemicellulose, xylose can consist up to 70−80% of the resulting hydrolysates (Lee and Jeffries, 2011). As xylose uptake by E. coli is significantly slower than glucose, optimization has been accomplished by strategies including genetic engineering (Dien et al, 2003), co-culturing (Eiteman et al, 2009), implementing non-native uptake pathways (Cam et al, 2016;Huo et al, 2017) ameliorating catabolite repression (Lu et al, 2016), or fine tuning growth conditions. One of the recent reports is to utilize xylose as the sole carbon source to produce γ-aminobutyric acid with a titer or 3.95 g/L (Zhao et al, 2017).…”

Process optimization, metabolic engineering interventions and commercialization of microbial polyhydroxyalkanoates production – A state‐of‐the art review

“…The recombinant strain produced a maximum of 21.0 g L −1 PHB from glucose and xylose. [ 137 ] Wang et al. [ 138 ] reported production of 400 mg L −1 mcl‐ PHA from recombinant E. coli .…”