Philipp, Laura-AlinaLaura-AlinaPhilippKneuer, LukasLukasKneuerMayer-Windhorst, CarinaCarinaMayer-WindhorstJautelat, SimonSimonJautelatLe, Nhat QuangNhat QuangLeGescher, JohannesJohannesGescher2025-07-092025-07-092025-06-01Applied and Environmental Microbiology 91 (6): e00685-25 (2025)https://hdl.handle.net/11420/56140Research in electro-microbiology provides unique opportunities to study and exploit microbial physiology. Several efforts have been made to transplant the extracellular electron transport chain from the native exoelectrogenic model organism Shewanella oneidensis into Escherichia coli. However, systematic comparisons between donor and recipient strain configurations are largely missing. Hence, the proposed minimal protein set, consisting of the c-type cytochromes cytoplasmic membrane protein A (CymA), small tetraheme cytochrome (STC), MtrA, and MtrC, as well as the β-barrel protein MtrB, was heterologously expressed in E. coli in different expansion stages. These stages were compared to corresponding S. oneidensis strains in terms of anthraquinone-2,6-disulfonate (AQDS) and ferric citrate reduction rates. This revealed that transplantation of heterologous extracellular electron transfer (EET) chains is associated with a tremendous decrease in electron transfer rates. As the acquired electron transfer rates were not competitive to S. oneidensis, it was hypothesized that protein localization and maturation might be affected by heterologous expression. Hence, the type II secretion system from S. oneidensis was also transplanted into an E. coli strain. The latter allowed the secretion of the terminal reductase MtrC onto the cell surface of E. coli for the first time. This was correlated with significantly increased but still insufficient extracellular electron transfer rates. Further experiments suggest that the correct folding of MtrB might be a further bottleneck.en1098-533620256American Society for Microbiologyhttps://creativecommons.org/licenses/by/4.0/c-type cytochromes | exoelectrogens | heterologous gene expression | microbiology | synthetic biologyNatural Sciences and Mathematics::570: Life Sciences, BiologyTechnology::616: DeseasesJournal Articlehttps://doi.org/10.15480/882.1536610.1128/aem.00685-2510.15480/882.15366Journal Article