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Research Focus:

Research in the Applied Biocatalysis group focusses on whole-cell biocatalysis and aims at the development of efficient and stable bioprocesses via an integrated biocatalyst and reaction engineering approach. Specifically, redox biotransformations (oxygenases, dehydrogenases, hydrogenases), in vivo cascades / orthogonal pathways, and fermentative approaches are targeted via enzyme, metabolic, reaction, and process engineering to develop and apply engineered microbial cells for the eco-efficient production of bulk and fine chemicals as well as fuels and bioactives from renewable as well as fossil resources. The renewables considered include sunlight, water and CO2  and thus the exploitation of photosynthesis with H2 formation by means of O2-tolerant hydrogenases as a major focus. Cyanobacteria, E. coli, and Pseudomonads constitute the host strains mainly employed. Following a systems biotechnology approach, special emphasis lies on biocatalyst and process efficiency with the ultimate goal to develop environment-friendly and industrially feasible processes via rational biocatalyst and process engineering.


Group Leader:
Prof. Dr. Bruno Bühler


Academic Staff:

Dr. Jörg Toepel

Dr. Adrian Tüllinghoff

Filipa Cantanhede


PhD-Students:

Lyn Katinka Möhrlin

Nolan Tucker


Bioprocess Engineer:

Daniel Spindler


Guest Scientist:

Carolin Bertelmann

Paul Bolay

Mahir Bozan

Sara Lupacchini

Ana Camila Zenteno Illanes

Nina Antonia Siebert


Students / Hiwis:

Mohidul Islam

Tizian Klarer

Index:

You could use our publication index for further requests.

2025 (2)

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2024 (5)

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2023 (3)

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2022 (6)

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2021 (6)

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2020 (5)

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2019 (8)

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2018 (1)

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2017 (6)

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2016 (2)

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2015 (5)

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Project: AI-supported biotechnology for resource-efficient active compound and bio-nylon production (BIOWIN)

(JTF InfraProNet 2021-2027, proposal-no. 100702818)

The project aims to develop innovative and sustainable technologies for the production of the antifibrinolytic agent ε-aminocaproic acid and bio-nylon monomers. The production strategies are based on the same artificial metabolic pathways, which will be optimized to maximize production rates. E. coli, Pseudomonads, and cyanobacterial strains are used or developed as production strains. Reaction technology and process concepts are being developed for these strains, including biofilm-based concepts. In order to accelerate process development, a paradigm shift from empirical to systematic approaches using AI and in-silico modeling will be implemented.


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Project: New Green Chemistry a contribution to a sustainable bioeconomy

(Proposal-no: 100328904)

The action New Green Chemistry extends the intrastructure at the Helmholtz Centre for Environmental Research GmbH – UFZ.

The "New Green Chemistry" approach aims at new processes, which consistently move ressources such as minerals and CO2 in circles, do not utilize agricultural areas and only cause minor climate-relevant emissions. Thereby, CO2 is assimilated by algae, but not to form biomass, but the organic acid glycolate. This acid is then used in a second process step in place of glucose and without harvesting and purification efforts for the microbial production of building blocks of the chemical industry. Prospectively, largely all products, which today are based on glucose (or starch) as carbon source, may be produced in this way.

The bioreactor equipment financed by this project will be utilized for the cultivation of heterotrophic microorganisms to develop the second process step and evaluate the coupling of both process steps.

Lecture: “Basics of Biocatalysis” in the 1st semester module “Introduction to Chemical Biotechnology” and "Drug target identification and validation"
Lecturer: Prof. Bruno Bühler

When:  Fridays in the winter semester, 1 pm – 3 pm or according to announcement
Where: MLU Halle, Weinberg-Campus, Hoher Weg 8, SR 101 HW or SR 107 HW according to announcement
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Lecture: “Whole-Cell Biocatalysis” in the 3rd semester module “Applied Biocatalysis”
Lecturer: Prof. Bruno Bühler

When:  Fridays in the winter semester, 10 am - 12 am or according to announcement
Where: MLU Halle, Weinberg-Campus, Hoher Weg 8, SR 101 HW or SR 107 HW according to announcement

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Project Seminar/ practical course on Applied Biocatalysis
Responsible Supervisor: Prof. Bruno Bühler

When:  One week in the winter semester break according to announcement
Where: UFZ, Leipzig

Master thesis 1:

Light-driven terminal hydroxylation of medium-chain alkanes using recombinant strains of Synechocystis sp.

supervisors: Dr. Adrian Tüllinghoff and Prof. Dr. Bruno Bühler

contact: adrian.tuellinghoff@ufz.de, bruno.buehler@ufz.de

Background: Terminal hydroxylation of medium-chain alkanes is a highly-desired enzymatic step for the production of polymer precursors from renewable fatty-acid methylesters (FAMEs).1 This oxygenation reaction can be conducted in a sustainable fashion when employing photosynthetically active microorganisms, such as cyanobacteria, as whole-cell biocatalysts. Coupling heterologous redox reactions to the photosynthetic light reaction in recombinant cyanobacteria gives access to light as energy and water as electron (and also O2) source.2 Previous studies showed that electron transfer from the photosynthetic apparatus to heterologous consumers, such as cytochrome P450 monooxygenases (CYP), is supported by host-intrinsic electron carriers providing electrons to the heterologous enzymes at high rates.3
Scope: Strains of the cyanobacterium Synechocystis sp. PCC 6803 expressing specific CYPs of the subfamily CYP153 are to be screened for terminal hydroxylation of hexanoic acid methyl ester (HAME). Natively, these CYPs are three-component systems, i.e., they consist of a NADH:ferredoxin-reductase (FNR), a ferredoxin (Fd), and the active oxygenase subunit. In Synechocystis sp. PCC 6803, a versatile set of host-intrinsic electron carriers was shown to (at least partly) take over the role of the native electron transfer partners FNR and Fd of a specific CYP from Acidovorax. By omitting FNR and/or Fd, it will be tested, if this compensation also occurs for the CYP153 enzymes used here. Next, new combinations of suitable Fd, FNR, and CYPs will be tested in vivo to optimize electron transfer towards product formation. The strain with optimized electron route will be tested in photobioreactors to examine feasibility at lab-scale.
Techniques:
- Biochemical techniques (activity assays, expression studies, CO spectra)
- Analytical techniques (HPLC, GC-FID)
- Microbiology techniques (Cultivation of phototrophic cells, transformation, microscopy)
- Bioprocess development (Feeding strategies, bioreactor cultivation and control)

Electron transfer routes from the photosynthetic apparatus to heterologous CYP-systems expressed in Synechocystis sp. PCC 6803:
Electron transfer routes from the photosynthetic apparatus to heterologous CYP-systems expressed in Synechocystis sp. PCC 6803.

References:
1. Ladkau, N., Assmann, M., Schrewe, M., Julsing, M. K., Schmid, A., and Bühler, B. (2016) Efficient production of the Nylon 12 monomer omega-aminododecanoic acid methyl ester from renewable dodecanoic acid methyl ester with engineered Escherichia coli, Metab Eng 36, 1-9.
2. Tüllinghoff, A., Djaya-Mbissam, H., Toepel, J., and Bühler, B. (2023) Light-driven redox biocatalysis on gram-scale in Synechocystis sp. PCC 6803 via an in vivo cascade, Plant Biotechnol J 21, 2074-2083.
3. Tüllinghoff, A., Toepel, J., and Bühler, B. (2024) Enlighting Electron Routes In Oxyfunctionalizing Synechocystis sp. PCC 6803, ChemBioChem 25, e202300475.


Master thesis 2:

Enhancing light-driven oxygenase biocatalysis by using membrane uncouplers for metabolic balancing

supervisors: Dr. Adrian Tüllinghoff and Prof. Dr. Bruno Bühler

contact: adrian.tuellinghoff@ufz.de, bruno.buehler@ufz.de

Background: To produce chemicals and goods in a truly sustainable and land-saving fashion, photo-biotechnology is a highly promising option.1 The photosynthetic apparatus of phototrophs enables the conversion of light into usable cellular energy. Cyanobacteria are ideal hosts for light-driven biocatalysis. Electron consuming biotransformations, such as oxygenase reactions, can be closely linked to the photosynthetic light reaction, giving access to light as energy and water as electron source.2, 3 The one-sided withdrawal of electron equivalents, such as NADPH, for heterologous redox reactions, however, is prone to affect photosynthetic metabolism.4 To reconstitute a more balanced ratio of ATP and NADPH levels, membrane uncouplers (uncoupling the proton gradient over the thylakoid membrane to reduce ATP formation) may help.
Scope: The impact of specific membrane uncouplers on the growth and the physiology of the model cyanobacterium Synechocystis sp. PCC 6803 is to be tested in different cultivation setups such as shaking flasks, bubble columns, and stirred-tank reactors. Further, options to enhance the biocatalytic performance of recombinant Synechocystis strains will be investigated, testing different uncoupler concentrations, time points of uncoupler supply and oxygenation rates. The findings at small scale will be transferred to photobioreactors at lab-scale to test the performance in an optimized production process. Depending on the progress, the project can be extended either by physiological characterization of the oxygenase-active biocatalyst or by testing downstream processing of the photo-biotechnological product.
Techniques:
- Biochemical techniques (activity assays, expression studies)
- Analytical techniques (HPLC, GC-FID, PAM fluorometry)
- Microbiology techniques (Cultivation of phototrophic cells, toxicity studies, microscopy)
- Bioprocess development (Feeding strategies, bioreactor cultivation and control)

Master thesis Uncoupler testing in Synechocystis
Light-driven production of 6-hydroxy hexanoic acid from cyclohexanone by a two-step enzyme cascade expressed in Synechocystis sp. PCC 6803.

References:

1. Brandenburg, F., Theodosiou, E., Bertelmann, C., Grund, M., Klähn, S., Schmid, A., and Krömer, J. O. (2021) Trans-4-hydroxy-L-proline production by the cyanobacterium Synechocystis sp. PCC 6803, Metab Eng Commun 12, e00155.
2. Hoschek, A., Toepel, J., Hochkeppel, A., Karande, R., Bühler, B., and Schmid, A. (2019) Light-dependent and aeration-independent gram-scale hydroxylation of cyclohexane to cyclohexanol by CYP450 harboring Synechocystis sp. PCC 6803, Biotechnol J 14, e1800724.
3. Tüllinghoff, A., Uhl, M. B., Nintzel, F. E., Schmid, A., Bühler, B., and Toepel, J. (2022) Maximizing photosynthesis-driven Baeyer-Villiger oxidation efficiency in recombinant Synechocystis sp. PCC 6803, Front Catal 1, art. 780474.
4. Tüllinghoff, A., Djaya-Mbissam, H., Toepel, J., and Bühler, B. (2023) Light-driven redox biocatalysis on gram-scale in Synechocystis sp. PCC 6803 via an in vivo cascade, Plant Biotechnol J 21, 2074-2083.