BioChem - DOS

Synthetically-primed adaptation of Pseudomonas putida to a non-native substrate D-xylose

Nature communications

Dvoř á k, Pavel; Burý š ková , Barbora; Popelá ř ová , Barbora; Ebert, Birgitta E.; Botka, Tibor; Bujdoš , Dalimil; Sá nchez-Pascuala, Alberto; Schö ttler, Hannah; Hayen, Heiko; de Lorenzo, Ví ctor; Blank, Lars M.; Beneš í k, Martin

<P><B>Abstract</B><P>To broaden the substrate scope of microbial cell factories towards renewable substrates, rational genetic interventions are often combined with adaptive laboratory evolution (ALE). However, comprehensive studies enabling a holistic understanding of adaptation processes primed by rational metabolic engineering remain scarce. The industrial workhorse <I>Pseudomonas putida</I> was engineered to utilize the non-native sugar D-xylose, but its assimilation into the bacterial biochemical network via the exogenous xylose isomerase pathway remained unresolved. Here, we elucidate the xylose metabolism and establish a foundation for further engineering followed by ALE. First, native glycolysis is derepressed by deleting the local transcriptional regulator gene <I>hexR</I>. We then enhance the pentose phosphate pathway by implanting exogenous transketolase and transaldolase into two lag-shortened strains and allow ALE to finetune the rewired metabolism. Subsequent multilevel analysis and reverse engineering provide detailed insights into the parallel paths of bacterial adaptation to the non-native carbon source, highlighting the enhanced expression of transaldolase and xylose isomerase along with derepressed glycolysis as key events during the process.</P></P>

2024

Springer Science and Business Media LLC

2041-1723

15

1

pp.2666

10.1038/s41467-024-46812-9

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