Experimental validation of in silico estimated biomass yields of Pseudomonas putida KT2440

Abstract

Pseudomonas putida is rapidly becoming a microbial cell platform for biotechnological applications. In order to understand genotype-phenotype relationships genome scale models represent helpful tools. However, the validation of in silico predictions of genome scale models is a task that is rarely performed. In this study the theoretical biomass yields of Pseudomonas putida KT2440 were estimated for 57 different carbon sources based on a genome scale stoichiometric model applying flux balance analysis. The batch growth of P. putida KT2440 with six individual carbon sources covering the range of maximal to minimal in silico biomass yields (acetate, glycerol, citrate, succinate, malate and methanol, respectively) was studied in a defined mineral medium in a fully controlled stirred-tank bioreactor on a 3 L scale. The highest growth rate of P. putida KT2440 was measured with succinate as carbon source (0.51 h ). Among the 57 carbon sources tested, glycerol resulted in the highest estimated biomass yield (0.61 molC molC ) which was experimentally confirmed. The comparison of experimental determined biomass yields with a modified version of the model iJP815 showed deviations of only up to 10%. The experimental data generated in this study can also be used in future studies to further improve the genome scale models of P. putida KT2440. Improved models will then help to gain deeper insights in genotype-phenotype relationships.

Pseudomonas putida KT2440 is naturally endowed to withstand industrial‐scale stress conditions

“…As depicted in Fig. 6, the theoretical 3‐HA‐based recovery of ATP can account for 79.6% of the cell’s ATP maintenance demand under glucose starvation assuming steady growth and applying the rather conservative P/O ratio of 1.33 (van Duuren et al , ; Hintermayer and Weuster‐Botz, ). However, if higher P/O ratios are considered, such as 1.75 (Nogales et al , ; Lieder et al , ; Beckers et al , ) or 1.85 (Hardy et al , ; Oberhardt et al , ), the total 3‐HA‐based recuperation of ATP would cover approximately 100% of ATP maintenance demand.…”

Biochemistry, genetics and biotechnology of glycerol utilization in Pseudomonas species

“…24% (Nikel et al., ). Hintermayer and Weuster‐Botz () simulated growth parameters of strain KT2440 in silico considering 57 individual carbon sources, and experimentally validated their prediction on six of them (acetate, glycerol, citrate, succinate, malate and CH 3 OH). Glycerol was found to promote the highest biomass yield on substrate (0.61 C‐mol C‐mol −1 ).…”

CRISPR interference‐mediated gene regulation in Pseudomonas putida KT2440

“…This strain has attracted substantial attention for industrial applications owing to its fast growth, native metabolic versatility and high robustness in harsh environmental conditions, which are particularly suitable traits for bio‐remediation of environmental contamination and metabolic engineering (Nikel et al ., ; Nikel and de Lorenzo, ). In addition, genome‐scale metabolic models of P. putida KT2440 were developed to better understand the versatile metabolism of the strain at the systems level, which were applied to reprogram the phenotype for biotechnological implementations (Nogales et al ., ; Sohn et al ., ; Hintermayer and Weuster‐Botz, ). Indeed, well‐established molecular biological tools, including systems for gene expression and genome engineering, have been harnessed in P. putida KT2440 to produce biofuels and pharmaceuticals by metabolic engineering (Martínez‐García and de Lorenzo, ; Cook et al ., ).…”