This title appears in the Scientific Report :
2019
Please use the identifier:
http://dx.doi.org/10.1186/s12934-019-1193-y in citations.
Please use the identifier: http://hdl.handle.net/2128/23068 in citations.
A FRET-based biosensor for the quantification of glucose in culture supernatants of mL scale microbial cultivations
A FRET-based biosensor for the quantification of glucose in culture supernatants of mL scale microbial cultivations
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 senso...
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Personal Name(s): | Otten, Julia |
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Tenhaef, Niklas / Jansen, Roman P. / Döbber, Johannes / Jungbluth, Lisa / Noack, Stephan / Oldiges, Marco / Wiechert, Wolfgang / Pohl, Martina (Corresponding author) | |
Contributing Institute: |
Biotechnologie; IBG-1 Strukturbiochemie; ICS-6 |
Published in: | Microbial cell factories, 18 (2019) 1, S. 143 |
Imprint: |
London
Biomed Central
2019
|
DOI: |
10.1186/s12934-019-1193-y |
PubMed ID: |
31434564 |
Document Type: |
Journal Article |
Research Program: |
Biotechnology |
Link: |
OpenAccess OpenAccess |
Publikationsportal JuSER |
Please use the identifier: http://hdl.handle.net/2128/23068 in citations.
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. |