Open AccessApplying systems biology tools to study n ‐butanol degradation in Pseudomonas putida KT2440
Author(s) -
Vallon Tobias,
Simon Oliver,
RendgenHeugle Beate,
Frana Sabine,
Mückschel Björn,
Broicher Alexander,
SiemannHerzberg Martin,
Pfannenstiel Jens,
Hauer Bernhard,
Huber Achim,
Breuer Michael,
Takors Ralf
Publication year - 2015
Publication title -
engineering in life sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.547
H-Index - 57
eISSN - 1618-2863
pISSN - 1618-0240
DOI - 10.1002/elsc.201400051
Subject(s) - pseudomonas putida , butanol , citric acid cycle , chemostat , biochemistry , metabolomics , metabolic engineering , n butanol , biology , metabolic flux analysis , pentose phosphate pathway , metabolism , tricarboxylic acid , chemistry , ethanol , bacteria , glycolysis , bioinformatics , enzyme , genetics
To smoothen the process of n ‐butanol formation in Pseudomonas putida KT2440, detailed knowledge of the impact of this organic solvent on cell physiology and regulation is of outmost importance. Here, we conducted a detailed systems biology study to elucidate cellular responses at the metabolic, proteomic, and transcriptional level. Pseudomonas putida KT2440 was cultivated in multiple chemostat fermentations using n ‐butanol either as sole carbon source or together with glucose. Pseudomonas putida KT2440 revealed maximum growth rates (μ) of 0.3 h −1 with n ‐butanol as sole carbon source and of 0.4 h −1 using equal C‐molar amounts of glucose and n ‐butanol. While C‐mole specific substrate consumption and biomass/substrate yields appeared equal at these growth conditions, the cellular physiology was found to be substantially different: adenylate energy charge levels of 0.85 were found when n‐ butanol served as sole carbon source (similar to glucose as sole carbon source), but were reduced to 0.4 when n ‐butanol was coconsumed at stable growth conditions. Furthermore, characteristic maintenance parameters changed with increasing n ‐butanol consumption. 13 C flux analysis revealed that central metabolism was split into a glucose‐fueled Entner–Doudoroff/pentose‐phosphate pathway and an n ‐butanol‐fueled tricarboxylic acid cycle when both substrates were coconsumed. With the help of transcriptome and proteome analysis, the degradation pathway of n ‐butanol could be unraveled, thus representing an important basis for rendering P. putida KT2440 from an n ‐butanol consumer to a producer in future metabolic engineering studies.