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@ARTICLE{Otten:865570,
author = {Otten, Julia and Tenhaef, Niklas and Jansen, Roman P. and
Döbber, Johannes and Jungbluth, Lisa and Noack, Stephan and
Oldiges, Marco and Wiechert, Wolfgang and Pohl, Martina},
title = {{A} {FRET}-based biosensor for the quantification of
glucose in culture supernatants of m{L} scale microbial
cultivations},
journal = {Microbial cell factories},
volume = {18},
number = {1},
issn = {1475-2859},
address = {London},
publisher = {Biomed Central},
reportid = {FZJ-2019-04935},
pages = {143},
year = {2019},
abstract = {BackgroundIn most microbial cultivations d-glucose is the
main carbon and energy source. However, quantification of
d-glucose especially in small scale is still challenging.
Therefore, we developed a FRET-based glucose biosensor,
which can be applied in microbioreactor-based cultivations.
This sensor consists of a glucose binding protein sandwiched
between two fluorescent proteins, constituting a FRET pair.
Upon d-glucose binding the sensor undergoes a conformational
change which is translated into a FRET-ratio
change.ResultsThe selected sensor shows an apparent Kd below
1.5 mM d-glucose and a very high sensitivity of up to $70\%$
FRET-ratio change between the unbound and the
glucose-saturated state. The soluble sensor was successfully
applied online to monitor the glucose concentration in an
Escherichia coli culture. Additionally, this sensor was
utilized in an at-line process for a Corynebacterium
glutamicum culture as an example for a process with
cell-specific background (e.g. autofluorescence) and
medium-induced quenching. Immobilization of the sensor via
HaloTag® enabled purification and covalent immobilization
in one step and increased the stability during application,
significantly.ConclusionA FRET-based glucose sensor was used
to quantify d-glucose consumption in microtiter plate based
cultivations. To the best of our knowledge, this is the
first method reported for online quantification of d-glucose
in microtiter plate based cultivations. In comparison to
d-glucose analysis via an enzymatic assay and HPLC, the
sensor performed equally well, but enabled much faster
measurements, which allowed to speed up microbial strain
development significantly.},
cin = {IBG-1 / ICS-6},
ddc = {570},
cid = {I:(DE-Juel1)IBG-1-20101118 / I:(DE-Juel1)ICS-6-20110106},
pnm = {581 - Biotechnology (POF3-581)},
pid = {G:(DE-HGF)POF3-581},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:31434564},
UT = {WOS:000483292300003},
doi = {10.1186/s12934-019-1193-y},
url = {https://juser.fz-juelich.de/record/865570},
}