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@PHDTHESIS{Lechtenberg:1034398,
author = {Lechtenberg, Thorsten},
title = {{T}olerance engineering of {P}seudomonas for the efficient
conversion and production of aldehydes},
volume = {292},
school = {Düsseldorf},
type = {Dissertation},
address = {Jülich},
publisher = {Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag},
reportid = {FZJ-2024-07180},
isbn = {978-3-95806-817-9},
series = {Schriften des Forschungszentrums Jülich Reihe
Schlüsseltechnologien / Key Technologies},
pages = {XVI, 185},
year = {2025},
note = {Dissertation, Düsseldorf, 2024},
abstract = {Biocatalysis holds promise to tackle the sustainability
challenges faced by chemical industry due to climate change
and depletion of fossil resources. However, obstacles emerge
regarding the compatibility of several important chemicals,
notably aldehydes, with biological systems, even if
remarkably robust workhorses such as bacteria of the
Pseudomonas clade are employed. This is related to the high
and versatile reactivity of aldehydes, which is both their
greatest asset and the root cause of their toxicity.
Competitive biocatalytic processes involving these
substances thus require tolerance-improved host organisms.
In view of the constantly growing demand for renewable and
ecologically produced plastics, the biocatalytic oxidation
of the burgeoning platform chemical
5-(hydroxymethyl)furfural (HMF) to 2,5-furandicarboxylic
acid (FDCA) is of particular interest since FDCA can
substitute structurally similar and fossilbased terephthalic
acid in polyesters. With the periplasmic oxidoreductase
complex PaoEFG and the cytoplasmic dehydrogenases AldB-I and
AldB-II, the primary enzymes responsible for the oxidation
of HMF and further aromatic aldehydes like
4-hydroxybenzaldehyde and vanillin by P. taiwanensis VLB120
and P. putida KT2440 were uncovered. This marks a
significant advancement from former black-box application of
these strains to specialized biocatalysts with fine-tuned
properties. To illustrate, overexpression of the newly
characterized genes resulted in so-called BOX-strains
(Boosted OXidation) with up to tenfold increased initial
oxidation rates in comparison to the wild type. As a result,
the new variants exhibited increased robustness when growing
in presence of HMF and also proved to be more efficient for
the complete oxidation of the aldehyde to the industrial
target compound FDCA. Furthermore, tolerance mechanisms
distinct from rapid oxidation were sought applying an
adaptive laboratory evolution approach. A ROX (Reduced
OXidation) deletion mutant with diminished aldehyde
conversion ability was subjected to steady HMF stress. This
yielded toleranceimproved strains through the unforeseen
inactivation of the regulator MexT and the associated
shutdown of the efflux pump MexEF-OprN. Another potential
use for oxidation-deficient, yet solvent-tolerant,
Pseudomonads is the biosynthesis of aromatic aldehydes, as
showcased with the popular aroma compound t-cinnamaldehyde.
In conclusion, this thesis contributes to the fundamental
understanding of aromatic aldehyde conversion by P.
taiwanensis VLB120 and P. putida KT2440 by unveiling the
underlying enzymes which were shown to constitute the
organisms’ main tolerance mechanism against these toxic
substances. Their overexpression in BOX strains strongly
increases aldehyde tolerance, and enables improved FDCA
production by boosted HMF oxidation. Reduced aldehyde
oxidation and reduction (ROAR) unlocks P. taiwanensis VLB120
for the (de novo) production of valuable aromatic aldehydes
or aldehyde-derived products, thereby expanding the product
portfolio of this aspiring microbial cell factory.},
cin = {IBG-1},
cid = {I:(DE-Juel1)IBG-1-20101118},
pnm = {2172 - Utilization of renewable carbon and energy sources
and engineering of ecosystem functions (POF4-217)},
pid = {G:(DE-HGF)POF4-2172},
typ = {PUB:(DE-HGF)3 / PUB:(DE-HGF)11},
urn = {urn:nbn:de:0001-2505050903420.294942610928},
doi = {10.34734/FZJ-2024-07180},
url = {https://juser.fz-juelich.de/record/1034398},
}