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@PHDTHESIS{Li:791657,
author = {Li, Wing Jin},
othercontributors = {Blank, Lars M. and Wierckx, Nick},
title = {{P}lastic monomer degradation - {E}ngineering {P}seudomonas
putida {KT}2440 for plastic monomer utilization; 1.
{A}uflage},
volume = {20},
school = {RWTH Aachen University},
type = {Dissertation},
address = {Aachen},
publisher = {Apprimus},
reportid = {RWTH-2020-05526},
isbn = {978-3-86359-858-7},
series = {Applied microbiology},
pages = {1 Online-Ressource (XVII, 162 Seiten) : Illustrationen,
Diagramme},
year = {2019},
note = {Auch veröffentlicht auf dem Publikationsserver der RWTH
Aachen University 2020; Dissertation, RWTH Aachen
University, 2019},
abstract = {Plastics are robust, ubiquitous, versatile materials, which
make our everyday life easier. But these polymers properties
make them both a blessing and a curse. The environmental
impact of plastics is immense, and so far, there are only a
few strategies to deal with plastic waste in an
environmentally friendly and economically feasible way. To
tackle this challenge, a strategy called bio-upcycling was
developed, aiming for a biotechnological conversion of
plastic waste like PET and PU. These polymers can be
hydrolysed by enzymes, releasing monomers like ethylene
glycol, 1,4-butanediol, and adipic acid. These can be
utilized as carbon source by microorganisms like the
biotechnological workhorse Pseudomonas putida KT2440 to
produce value-added compounds. With this strategy, plastic
waste can be used to produce value-added materials. This
thesis aims to enable P. putida KT2440 to efficiently
metabolize the plastic monomers ethylene glycol,
1,4-butanediol, and adipic acid. Since P. putida KT2440 is
not able to grow on ethylene glycol, adaptive laboratory
evolution (ALE) was performed to isolate the enhanced
mutants. Genome resequencing and reverse engineering
revealed that the deletion of one regulator, gclR, was
sufficient to enable growth on EG. The deletion of two
additionally identified genes, $PP_2046$ and $PP_2662,$
could further enhance this growth. With this knowledge,
ethylene glycol metabolism and its regulation in P. putida
was further unraveled. A similar ALE-based strategy was
applied to enhance growth on 1,4-butanediol and to enlighten
underlying degradation pathways. Targets like $PP_2046$ were
identified via genome resequencing, and proteomic analysis
gave insights to the involvement of dehydrogenases. With the
overexpression of the operon $PP_2047-51,$ higher growth
rates were achieved. Further characterizations indicate that
1,4-butanediol is at least in part metabolized through
β-oxidation. Adipic acid metabolism was enabled in P.
putida by introducing the heterologous genes dcaAKIJP from
Acinetobacter baylyi and subsequent ALE. Furthermore, genome
resequencing analysis suggests a hybrid adipic acid
metabolic pathway involving DcaAKIJP from A. baylyi combined
with parts of native phenylacetate degradation, and
β-oxidation pathways. In addition to enabling P. putida
KT2440 to grow on these substrates individually, a strain
was also engineered to metabolize all three of these
plastic-derived compounds. Thus, this work sets the basis
for bio-upcycling of PET and PU and leads the way for the
rational design of a consolidated plastic degrader.},
cin = {161710 / 160000},
ddc = {570},
cid = {$I:(DE-82)161710_20140620$ / $I:(DE-82)160000_20140620$},
typ = {PUB:(DE-HGF)11 / PUB:(DE-HGF)3},
doi = {10.18154/RWTH-2020-05526},
url = {https://publications.rwth-aachen.de/record/791657},
}
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