Recombinant Ralstonia eutropha engineered to utilize xylose and its use for the production of poly(3-hydroxybutyrate) from sunflower stalk hydrolysate solution

Abstract

BackgroundLignocellulosic raw materials have extensively been examined for the production of bio-based fuels, chemicals, and polymers using microbial platforms. Since xylose is one of the major components of the hydrolyzed lignocelluloses, it is being considered a promising substrate in lignocelluloses based fermentation process. Ralstonia eutropha, one of the most powerful and natural producers of polyhydroxyalkanoates (PHAs), has extensively been examined for the production of bio-based chemicals, fuels, and polymers. However, to the best of our knowledge, lignocellulosic feedstock has not been employed for R. eutropha probably due to its narrow spectrum of substrate utilization. Thus, R. eutropha engineered to utilize xylose should be useful in the development of microbial process for bio-based products from lignocellulosic feedstock.ResultsRecombinant R. eutropha NCIMB11599 expressing the E. coli xylAB genes encoding xylose isomerase and xylulokinase respectively, was constructed and examined for the synthesis of poly(3-hydroxybutyrate) [P(3HB)] using xylose as a sole carbon source. It could produce 2.31 g/L of P(3HB) with a P(3HB) content of 30.95 wt% when it was cultured in a nitrogen limited chemically defined medium containing 20.18 g/L of xylose in a batch fermentation. Also, recombinant R. eutropha NCIMB11599 expressing the E. coli xylAB genes produced 5.71 g/L of P(3HB) with a P(3HB) content of 78.11 wt% from a mixture of 10.05 g/L of glucose and 10.91 g/L of xylose in the same culture condition. The P(3HB) concentration and content could be increased to 8.79 g/L and 88.69 wt%, respectively, when it was cultured in the medium containing 16.74 g/L of glucose and 6.15 g/L of xylose. Further examination of recombinant R. eutropha NCIMB11599 expressing the E. coli xylAB genes by fed-batch fermentation resulted in the production of 33.70 g/L of P(3HB) in 108 h with a P(3HB) content of 79.02 wt%. The concentration of xylose could be maintained as high as 6 g/L, which is similar to the initial concentration of xylose during the fed-batch fermentation suggesting that xylose consumption is not inhibited during fermentation. Finally, recombinant R. eutorpha NCIMB11599 expressing the E. coli xylAB gene was examined for the production of P(3HB) from the hydrolysate solution of sunflower stalk. The hydrolysate solution of sunflower stalk was prepared as a model lignocellulosic biomass, which contains 78.8 g/L of glucose, 26.9 g/L of xylose, and small amount of 4.8 g/L of galactose and mannose. When recombinant R. eutropha NCIMB11599 expressing the E. coli xylAB genes was cultured in a nitrogen limited chemically defined medium containing 23.1 g/L of hydrolysate solution of sunflower stalk, which corresponds to 16.8 g/L of glucose and 5.9 g/L of xylose, it completely consumed glucose and xylose in the sunflower stalk based medium resulting in the production of 7.86 g/L of P(3HB) with a P(3HB) content of 72.53 wt%.ConclusionsRalstonia eutropha was successfully engineered to utilize xylose as a sole carbon source as w...

Improving polyhydroxyalkanoate production from inexpensive carbon sources by genetic approaches: a review

“…The recombinant strains produced limited PHB amounts from arabinose or xylose but these studies provide the basis for the development of C. necator strains with PHA‐producing ability from pentose sugars, which are available in lignocellulosic hydrolysates. A noteworthy paper recently reported obtaining high P(3HB) content (73% CDM) from sunflower stalk hydrolysate by means of engineered R. eutropha pKM212‐XylAB expressing the E. coli xylAB genes encoding xylose isomerase and xylulokinase, thus efficiently co‐utilizing glucose and xylose …”

“…A noteworthy paper recently reported obtaining high P(3HB) content (73% CDM) from sunflower stalk hydrolysate by means of engineered R. eutropha pKM212-XylAB expressing the E. coli xylAB genes encoding xylose isomerase and xylulokinase, thus efficiently co-utilizing glucose and xylose. 94…”

Construction of a Vitreoscilla Hemoglobin Promoter-Based Tunable Expression System for Corynebacterium glutamicum

“…Biorefinery processes have been developed to establish a sustainable alternative to produce chemicals, polymers, and fuels, and they currently rely on petroleum-based processes [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17]. Development of recombinant microorganisms capable of converting a broader range of renewable biomass feedstock into bio-based chemicals, with properties comparable to those of conventional petrochemical products, have been extensively studied [18][19][20][21][22][23][24][25][26][27]. For example, processes for the production of bio-polymers, such as polylactic acid (PLA) and polybutylene succinate (PBS) in a biorefinery have been established [21][22][23].…”

Metabolic engineering of Corynebacterium glutamicum for enhanced production of 5-aminovaleric acid

“…As a result of increasing pressure on the environment, bio-based production of chemicals, fuels, and materials from renewable non-food biomasses has been attracting much attention [ 1 ]. To make such bio-based processes competitive, microorganisms have been metabolically engineered for production of fuels [ 2 – 4 ], amino acids [ 5 – 9 ], polymers [ 10 – 12 ], and other chemicals of industrial importance [ 13 – 15 ]. It is expected that more chemicals and materials of petrochemical origin will be produced through bio-based route employing microorganisms developed by systems metabolic engineering [ 16 , 17 ].…”