Energy converting membrane enzymes in fatty acid degrading, syntrophic and pathogenic bacteria

• Applicant: Professor Dr. Matthias Boll
• Subject Area: Metabolism, Biochemistry and Genetics of Microorganisms ; Biochemistry
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 560368629

The microbial conversion of natural polymers into methane plays a significant role in the global carbon cycle, accounting for over half of all methane produced on Earth annually. In this process, anaerobic syntrophic bacteria convert the products of primary fermenting bacteria, including short-chain fatty acids, into hydrogen gas, formate, and acetate. These secondary fermentation products then serve as substrate for methanogenic archaea. We have recently identified an energy-converting membrane protein that plays a pivotal role in this process: the electron-transferring flavoprotein (ETF):methylmenaquinone (MMK) oxidoreductase (EMO). It is predicted that this enzyme, in conjunction with a membrane-bound formate dehydrogenase, will facilitate the endergonic electron transfer from acyl-CoA to CO₂ via a reverse, proton motive force-dependent redox-loop during the beta-oxidation of fatty acids. MMK serves as membrane-bound electron carrier. An unconventional ATP synthase has the sole function of generating a proton motive force and is predicted to transport four protons outside the cell per ATP hydrolyzed. EMOs are present in a wide range of (M)MK-containing bacteria with the capability of fatty acid beta-oxidation, including Mycobacterium tuberculosis. In this pathogen, EMO is believed to be a vital factor in the ability to persist within macrophages. The present project is guided by three objectives. The objective 1 addresses to the structural, spectroscopic and functional characterization of EMO from the syntrophic Deltaproteobacterium Syntrophus aciditrophicus. It will be accomplished through the utilization of cryo-electron microscopy and electron paramagnetic resonance spectroscopy on both wild-type enzymes and molecular variants. In order to elucidate the function of EMO in the predicted redox reverse loop, the enzyme will be reconstituted in proteoliposomes or inside-out vesicles derived from S. aciditrophicus cells. (ii) The objective 2 is to characterize the structure and function of EMO from M. tuberculosis, which represents an attractive target for novel antimycobacterial drugs. (iii) Objective 3 is to provide the first structural and functional characterization of an ATP synthase from a syntrophic organism and to present experimental evidence for the predicted proton-pumping/ATP hydrolysis stoichiometry. The anticipated outcome of this project is to gain a comprehensive understanding of the bioenergetics of syntrophic scFAs oxidation, a pivotal process in the global carbon cycle and biogas formation.

• DFG Programme: Research Grants