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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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Record number 432030
Title In-silico-driven metabolic engineering of Pseudomonas putida for enhanced production of poly-hydroxyalkanoates
Author(s) Poblete-Castro, I.; Binger, D.; Rodrigues, A.; Becker, J.; Martins Dos Santos, V.A.P.; Wittmann, C.
Source Metabolic Engineering 15 (2013). - ISSN 1096-7176 - p. 113 - 123.
DOI http://dx.doi.org/10.1016/j.ymben.2012.10.004
Department(s) Systems and Synthetic Biology
VLAG
Publication type Refereed Article in a scientific journal
Publication year 2013
Keyword(s) chain-length polyhydroxyalkanoates - gram-negative bacteria - fed-batch culture - genome sequence - escherichia-coli - kt2440 - pathways - acid - poly(3-hydroxyalkanoates) - biosynthesis
Abstract Here, we present systems metabolic engineering driven by in-silico modeling to tailor Pseudomonas putida for synthesis of medium chain length PHAs on glucose. Using physiological properties of the parent wild type as constraints, elementary flux mode analysis of a large-scale model of the metabolism of P. putida was used to predict genetic targets for strain engineering. Among a set of priority ranked targets, glucose dehydrogenase (encoded by gcd) was predicted as most promising deletion target. The mutant P. putida ¿gcd, generated on basis of the computational design, exhibited 100% increased PHA accumulation as compared to the parent wild type, maintained a high specific growth rate and exhibited an almost unaffected gene expression profile, which excluded detrimental side effects of the modification. A second mutant strain, P. putida ¿pgl, that lacked 6-phosphogluconolactonase, exhibited a substantially decreased PHA synthesis, as was also predicted by the model. The production potential of P. putida ¿gcd was assessed in batch bioreactors. The novel strain showed an increase of the PHA yield (+80%), the PHA titer (+100%) and cellular PHA content (+50%) and revealed almost unaffected growth and diminished by-product formation. It was thus found superior in all relevant criteria towards industrial production. Beyond the contribution to more efficient PHA production processes at reduced costs that might replace petrochemical plastics in the future, the study illustrates the power of computational prediction to tailor microbial strains for enhanced biosynthesis of added-value compounds
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