Heterologous rhamnolipid biosynthesis by P. putida KT2440 on bio-oil derived small organic acids and fractions

Abstract

In many cases in industrial biotechnology, substrate costs make up a major part of the overall production costs. One strategy to achieve more cost-efficient processes in general is to exploit cheaper sources of substrate. Small organic acids derived from fast pyrolysis of lignocellulosic biomass represent a significant proportion of microbially accessible carbon in bio-oil. However, using bio-oil for microbial cultivation is a highly challenging task due to its strong adverse effects on microbial growth as well as its complex composition. In this study, the suitability of bio-oil as a substrate for industrial biotechnology was investigated with special focus on organic acids. For this purpose, using the example of the genetically engineered, non-pathogenic bacterium Pseudomonas putida KT2440 producing mono-rhamnolipids, cultivation on small organic acids derived from fast pyrolysis of lignocellulosic biomass, as well as on bio-oil fractions, was investigated and evaluated. As biosurfactants, rhamnolipids represent a potential bulk product of industrial biotechnology where substitution of traditional carbon sources is of conceivable interest. Results suggest that maximum achievable productivities as well as substrate-to-biomass yields are in a comparable range for glucose, acetate, as well as the mixture of acetate, formate and propionate. Similar yields were obtained for a pretreated bio-oil fraction, which was used as reference real raw material, although with significantly lower titers. As such, the reported process constitutes a proof-of-principle for using bio-oil as a potential cost-effective alternative carbon source in a future bio-based economy.

Killing Two Birds With One Stone – Strain Engineering Facilitates the Development of a Unique Rhamnolipid Production Process

“…A concentration of over 10 g L −1 acetate likely caused growth inhibition. Recently, Arnold et al (2019) tested 10 g L −1 acetate as the sole carbon source for rhamnolipid production with a recombinant P. putida KT2440 and also identified a growth inhibition. Further, an acetate accumulation during growth on ethanol was reported for P. putida KT2440 (Yang et al, 2019).…”

“…The overall goal is to establish a circular bioeconomy in which also lignocellulosic biomass and side- and waste streams are used as energy and carbon source. Several studies deal with the production of rhamnolipids from alternative resources, e.g., butane, agro-industrial waste, glycerol, organic acids derived from bio-oil, and xylose ( Gehring et al, 2016 ; Gudiña et al, 2016 ; Tiso et al, 2017 ; Arnold et al, 2019 ; Bator et al, 2020 ). In the 1990s, ethanol was already considered as an alternative carbon source for rhamnolipid production ( Matsufuji et al, 1997 ).…”

Heterologous Rhamnolipid Biosynthesis: Advantages, Challenges, and the Opportunity to Produce Tailor-Made Rhamnolipids

“…Recent studies are more focused on establishing unusual carbon sources or cultivation conditions to make the rhamnolipid biosynthesis more economical, e.g., by using cheap raw materials from waste streams. These studies were based on well-established host organisms like P. putida KT2440 engineered to utilize ethanol, pyrolysis oil or alternative sugars like xylose and arabinose as part of lignocellulosic hydrolysates or from agricultural residues (Arnold et al, 2019;Horlamus et al, 2019;Wang et al, 2019;Bator et al, 2020a,b) or they were cultivated in biofilms to avoid foaming as in conventional bioreactors (Wigneswaran et al, 2016). Moreover, new heterologous hosts for rhamolipid production were exploited, e.g., Cellvibrio japonicus (Horlamus et al, 2018), which can even utilize polymeric substrates like xylan and cellulose (Gardner, 2016), or Pseudomonas stutzeri, which was used to produce rhamnolipids under anaerobic conditions (Zhao et al, 2015; Table 1).…”

Coupling an Electroactive Pseudomonas putida KT2440 with Bioelectrochemical Rhamnolipid Production

“…P. aeruginosa is a facultative pathogen and its rhamnolipid production is controlled by a complex quorum sensing network [ 5 , 6 , 7 ]. Hence, research in the past years has focused on heterologous production with the non-pathogenic P. putida KT2440 [ 8 , 9 , 10 ].…”