Engineering xylose metabolism for production of polyhydroxybutyrate in the non-model bacterium Burkholderia sacchari

Abstract

BackgroundDespite its ability to grow and produce high-value molecules using renewable carbon sources, two main factors must be improved to use Burkholderia sacchari as a chassis for bioproduction at an industrial scale: first, the lack of molecular tools to engineer this organism and second, the inherently slow growth rate and poly-3-hydroxybutyrate [P(3HB)] production using xylose. In this work, we have addressed both factors.ResultsFirst, we adapted a set of BglBrick plasmids and showed tunable expression in B. sacchari. Finally, we assessed growth rate and P(3HB) production through overexpression of xylose transporters, catabolic or regulatory genes. Overexpression of xylR significantly improved growth rate (55.5% improvement), polymer yield (77.27% improvement), and resulted in 71% of cell dry weight as P(3HB).ConclusionsThese values are unprecedented for P(3HB) accumulation using xylose as a sole carbon source and highlight the importance of precise expression control for improving utilization of hemicellulosic sugars in B. sacchari.

Engineering the Osmotic State of Pseudomonas putida KT2440 for Efficient Cell Disruption and Downstream Processing of Poly(3-Hydroxyalkanoates)

“…Firstly, it was evaluated whether inactivation of the mscL gene and overexpression of the porin genes exerted a detrimental effect on bacterial growth of all constructed strains. Frequently basal transcriptional leakage of plasmidial genes or gene suppression generates a diminished biomass synthesis that will lastly affect PHA accumulation (Guamán et al, 2018). The growth rate of the genetically modified KT2440 strains, namely KT-oprF, KT-oprE, and KT-oprFE did not vary significantly compared with KT2440 wild-type using decanoate (20 mM) as the sole carbon source in minimal salt medium (Figure 2A).…”

Valorisation of xylose to renewable fuels and chemicals, an essential step in augmenting the commercial viability of lignocellulosic biorefineries

“…Polyhydroxyalkanoates (PHA) are the hydroxy alkanoic polyesters which are stored as intracellular granules in various prokaryotic microorganisms and are accumulated when carbon source is in surplus along with limitation of a key nutrient. 170,171 Although the primary function of these polyesters are the storage of carbon and energy, they also play a role in preventing the microbial cell from stress. Poly-3-hydroxybutyrate (PHB)…”

“…The engineered B. sacchari strain showed 55, 77.3 and 71% improvement in the growth rate, polymer yield and cell dry weight, respectively. 170 As explained in section 5.3.,…”

Desenvolvimentos de ferramentas matemáticas para modelagem de sistemas biológicos baseados em fluxos metabólicos e estimação de parâmetros de problemas mal condicionados.

“…However, non-model organisms may have distinct properties that are worth exploring (RUSSELL et al, 2017). For instance, Burkholderia sacchari is a non-model bacteria isolated from sugarcane crops in Brazil that has the potential for producing high-value molecules from renewable carbon sources with five or six carbon atoms (GUAMÁN et al, 2018).…”

“…However, when working with non-model organisms, those that have not been extensively studied, there is usually not sufficient information to apply these highly detailed models based on the complete genome. Despite that obstacle, non-model organisms are worth investigating, as they can reveal unique properties and have potential economical interest for industrial processes (GUAMÁN et al, 2018;ARMENGAUD et al, 2014). Taking it into consideration, the first part of this thesis describes the implementation of stoichiometric modeling techniques for small and medium metabolic networks that can help elucidate the physiological state of microorganisms and identify means to improve their metabolism to hopefully make their application in biotechnological processes viable.…”