S. (Sandy) Schmidt, Dr
Our research exploits the powerful reactivity and selectivity of enzymes from secondary metabolite pathways for the production of natural products and their analogs for pharmaceutical applications. One of the main areas of interest is the identification, characterization and the engineering of Rieske non-heme iron oxygenases for their application in biocatalytic reactions towards pharmaceuticals and/or precursors thereof. Along this line, the outstanding reactivities of these biocatalysts are exploited in synthetic metabolic pathways and (chemo)enzymatic cascade reactions for the production of complex compounds from simple precursors. The Schmidt group is thereby using state-of-the-art synthetic biology tools for the design and assembly of artificial metabolic pathways, and is developing new methodologies for the genetic engineering of several chassis strains, including autotrophic bacteria such as Cupriavidus necator. Finally, the Schmidt group aims at the development of new concepts for electron transfer pathways in microorganisms, and is developing concepts to optimize electron transfer chains in multi-component oxidoreductases. By expanding our knowledge on the parameters determining efficient electron transfer and thus catalysis, it is expected that this will increase the broad applicability of multi-component oxidoreductases relying on complex electron transfer mechanisms.
We are currently working on the following research topics:
A group of enzymes called Rieske non-heme iron oxygenases (ROs) catalyzes reactions that are among the most challenging in organic syntheses. ROs are the only enzymes known to catalyze the stereoselective formation of vicinal cis-diols in one step leading to important building blocks for pharmaceuticals. These enzymes are soluble multicomponent systems that harness the reductive power of NAD(P)H for oxygen activation. Due to their versatility, ROs are considered as the non-heme analogue of cytochrome P450 monooxygenases and, in addition to their relaxed substrate specificity, these enzymes can catalyze various oxidation reactions, resulting in an enormous potential of these enzymes for manifold synthetically useful transformations. In contrast to many flavin-dependent monooxygenases, ROs depend on electron transfer from a reductase or the interplay of a reductase (FdR) and a ferredoxin (Fd). In the latter case, the cofactor-derived electrons are transferred via FdR and Fd to the terminal oxygenase (Oxy) component containing the active site.
In the last decade, Rieske oxygenases have also been revealed to play important roles in natural product biosynthesis. Often, the pharmaceutical potential of natural product scaffolds is underexplored due to the difficulty in accessing analogs to complex secondary metabolites using traditional synthetic methods. Recent advances in sequencing technology have facilitated the identification of secondary metabolite gene clusters from genomic and metagenomic samples, which has led to an explosion of newly identified and annotated secondary metabolite pathways.
We strive to leverage the powerful reactivity and selectivity of ROs from natural product pathways in concise approaches to natural products and their analogs. We are particularly interested in the biosynthesis of hapalindole-type alkaloids that have gained increasing attention as highly potential leads in recent drug discovery due to their diverse bioactivities. We strive to identify putative ROs from the hapalindole-type alkaloids biosynthetic gene clusters and to elucidate their catalytic role in the synthesis and diversification of these natural products. We employ mechanistic studies with biochemical characterization along with the determination of the substrate scope for their application in synthesizing biologically active molecules. Thereby, synthetic efforts feed biological studies on activity, which inform the subsequent selection of synthetic targets.
Electron transfer pathways in enzymes
Rieske oxygenases depend on electron transfer from a reductase or the interplay of a reductase and a ferredoxin. These complex redox machines require multistep electron tunneling architectures that can transfer electrons rapidly with only a small loss of free energy to the surface of the oxygenase, where the actual catalysis takes place.
The applicability of enzymes depending on an electron transfer chain involving ferredoxins remains to be challenging, mainly due to their low catalytic activity, low stability, and dependence on a complex electron transfer system. Especially in the case of P450 monooxygenases, many efforts have been made to surmount these limitations by simplifying the electron transfer chain. This has led to increased knowledge on the electron transfer pathways in oxidoreductases, however, in Rieske oxygenases the underlying molecular mechanisms that determine the interactions between ferredoxins and their oxygenases are much less understood.
We strive to elucidate the mechanisms that determine the redox partner specificity in Rieske oxygenases and in turn to increase the understanding of the essential aspects determining an efficient electron transfer based on specific protein-protein interactions between the respective partners. We investigate to which extent the redox partner specificity in Rieske oxygneases can be altered toward non-natural electron donors, and whether electron transfer chains in these complex redox machines can be simplified to increase the applicability of ROs.
Photo-biocatalytic Cascades: The combination of several catalytic steps to conduct a precisely arranged sequence of chemical transformations in a single reaction vessel exhibits an enormous potential for more economically and ecologically efficient synthetic routes, and thus, the development of one-pot (cascade) reactions is a growing research field. Combining catalysts from different catalysis fields (“worlds”), however, is sometimes more challenging for compatibility reasons. Especially photocatalysts have been combined with various types of other catalysts yielding synergistic dual catalytic systems, where two types of catalysts participate in one catalytic cycle. The combination of photocatalytic and biocatalytic steps for organic synthesis, however, has not been systematically explored until now. Most examples of combining photo- and biocatalysis focus on photocatalytic in situ regeneration of redox enzymes.
We strive to couple photocatalytic reactions that use light energy directly to drive small molecule conversions with further enzymatic functionalization to develop photobiocatalytic concurrent tandem or sequential reactions. We investigate limitations associated to photochemoenzymatic cascades, and develop strategies to overcome the often observed incompatibilities between enzymes and photocatalysts.
H2020 MSCA ITN-EJD 764920 (PhotoBioCat)
Autotrophic chassis strain development
Chemists already strive for more than 100 years for possibilities to produce valuable chemicals from CO2. With the growing world population, depletion of fossil resources and the on-going climate change, the last years’ research strongly focuses on innovative and applicable solutions via chemical as well as biotechnological routes. Thereby, biotechnological routes provide several advantages over chemical strategies. With their capacity to utilize renewable energy for the accumulation of biomass from CO2, autotrophic microorganisms have an enormous potential for the selective production of a broad palette of future materials for our society.
However, the biotechnological utilization of CO2 is associated to several challenges, mainly related to the enzymatic fixation of CO2 to enter metabolism as well as the supply of energy, which is at the same time the greatest chance for microbial biotechnology in this context. In order to overcome these challenges, synthetic biology and process engineering must be integrated to demonstrate the successful production of commodities from CO2.
We will explore the potential of the autotrophic bacterium Cupriavidus necator for the production of biochemicals. Particular interest lies thereby on the combination of synthetic biology approaches with metabolic engineering for an adaptive laboratory evolution to create an efficient route from CO2 fixation to the production of commodities. The native machinery for H2/CO2-utilization in the CO2-fixing C. necator is highly optimized from evolution, and thus represents a tremendous space to connect the metabolism with additional electron sinks and to increase space-time yields. Based on this, we strive to develop robust C. necator production strains and investigate metabolic routes using the strengths of C. necator. In order to provide optimal metabolic supply for the envisioned products, we will develop molecular genetic tools for an efficient chassis strain engineering to produce several commodities from CO2.
H2020 MSCA ITN-EJD 955740 (ConCO2rde)
|Last modified:||25 June 2022 5.07 p.m.|