Structural and functional characterization of FccABC, a flavocytochrome c complex from Wolinella succinogenes

Abstract: The use of plastics has increased tremendously in every sector of life. The production of plastics has been increasing exponentially in the last 50 years. Thus managing the excess plastic waste has become a pressing environmental issue. This study focuses on a particular plastic molecule, methacrylate. So far, Geobacter sulfurreducens AM-1 is the only known bacterium capable of reducing methacrylate. The Flavocytochrome c flavin subunit, FccA, belonging to Wollinella succinogenes shares around 83 % sequence similarities with the methacrylate reductase from G. sulfurreducens AM-1. Also, FccA displays amino acid sequence homologies to flavocytochrome c fumarate reductases and to the catalytic subunit of the membrane-bound succinate: quinone oxidoreductase (respiratory complex II). Interestingly, FccA does not reduce fumarate. It is co-transcribed with FccB (a 14 kDa protein with homologies to the tetraheme domain of fumarate reductases) and FccC (a 24 kDa membrane-anchored tetraheme c-type cytochrome with homologies to the NirT/NapC family).
In this work, FccA, FccB, FccC, FccAB, FccBC, and FccABC, were heterologously expressed in E. coli, isolated and purified for structural and functional investigations. The catalytic protein, FccA was most extensively studied. Methacrylate reductase activity of FccA was confirmed using benzyl viologen linked reductive assays, making it the second known methacrylate reductase to date. The product of the assay was also confirmed to be iso-butyrate using ion chromatography. However, FccA was not able to catalyze the reduction of fumarate. The three-dimensional structure of FccA with and without bound methacrylate was determined by X-ray crystallography. This is the first known structure of methacrylate reductase available till now. The rest of the study focuses on the structural similarities between W. succinogenes FccA and S. putrefaciens fumarate reductase. In order to catalyze fumarate reduction by FccA, amino acid substitutions were undertaken in FccA active site mimicking the fumarate reductase. Along with the additional study performed on other subunits of the FccABC complex, a putative function of the FccABC complex is discussed focusing on the role of each protein

Object Details

• Location: Deutsche Nationalbibliothek Frankfurt am Main
• Extent: Online-Ressource
• Language: Englisch
• Notes: Universität Freiburg, Dissertation, 2020

Contributors, Places and Time

• Event: Veröffentlichung
• (where): Freiburg
• (who): Universität
• (when): 2020
• Creator: Dabhade, Prachi
• Contributor: Andrade, Susana ; Albert-Ludwigs-Universität Freiburg. Fakultät für Chemie und Pharmazie

Further information

• DOI: 10.6094/UNIFR/174561
• URN: urn:nbn:de:bsz:25-freidok-1745617
• Rights: Open Access; Der Zugriff auf das Objekt ist unbeschränkt möglich.
• Last update: 25.03.2025, 1:44 PM CET