@article{Rasmussen2023, author = {Rasmussen, Tim}, title = {The potassium efflux system Kef: bacterial protection against toxic electrophilic compounds}, series = {Membranes}, volume = {13}, journal = {Membranes}, number = {5}, issn = {2077-0375}, doi = {10.3390/membranes13050465}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-313686}, year = {2023}, abstract = {Kef couples the potassium efflux with proton influx in gram-negative bacteria. The resulting acidification of the cytosol efficiently prevents the killing of the bacteria by reactive electrophilic compounds. While other degradation pathways for electrophiles exist, Kef is a short-term response that is crucial for survival. It requires tight regulation since its activation comes with the burden of disturbed homeostasis. Electrophiles, entering the cell, react spontaneously or catalytically with glutathione, which is present at high concentrations in the cytosol. The resulting glutathione conjugates bind to the cytosolic regulatory domain of Kef and trigger activation while the binding of glutathione keeps the system closed. Furthermore, nucleotides can bind to this domain for stabilization or inhibition. The binding of an additional ancillary subunit, called KefF or KefG, to the cytosolic domain is required for full activation. The regulatory domain is termed K+ transport-nucleotide binding (KTN) or regulator of potassium conductance (RCK) domain, and it is also found in potassium uptake systems or channels in other oligomeric arrangements. Bacterial RosB-like transporters and K+ efflux antiporters (KEA) of plants are homologs of Kef but fulfill different functions. In summary, Kef provides an interesting and well-studied example of a highly regulated bacterial transport system.}, language = {en} } @phdthesis{Ibrahim2024, author = {Ibrahim, Eslam Samir Ragab}, title = {Unraveling the function of the old yellow enzyme OfrA in \(Staphylococcus\) \(aureus\) stress response}, doi = {10.25972/OPUS-28960}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-289600}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2024}, abstract = {Biological systems are in dynamic interaction. Many responses reside in the core concepts of biological systems interplay (competition and cooperation). In infection situation, the competition between a bacterial system and a host is shaped by many stressors at spatial and temporal determinants. Reactive chemical species are universal stressors against all biological systems since they potentially damage the basic requirements of these systems (nucleic acids, proteins, carbohydrates, and lipids). Either produced endogenously or exogenously, reactive chemical species affect the survival of pathogens including the gram-positive Staphylococcus aureus (S. aureus). Therefore, bacteria developed strategies to overcome the toxicity of reactive species. S. aureus is a widely found opportunistic pathogen. In its niche, S. aureus is in permanent contact with surrounding microbes and host factors. Deciphering the deterministic factors in these interactions could facilitate pinpointing novel bacterial targets. Identifying the aforementioned targets is crucial to develop new strategies not only to kill the pathogenic organisms but also to enhance the normal flora to minimize the pathogenicity and virulence of potential pathogens. Moreover, targeting S. aureus stress response can be used to overcome bacterial resistance against host-derived factors. In this study, I identify a novel S. aureus stress response factor against reactive electrophilic, oxygen, and hypochlorite species to better understand its resilience as a pathogen. Although bacterial stress response is an active research field, gene function is a current bottleneck in characterizing the understudied bacterial strategies to mediate stress conditions. I aimed at understanding the function of a novel protein family integrated in many defense systems of several biological systems. In bacteria, fungi, and plants, old yellow enzymes (OYEs) are widely found. Since the first isolation of the yellow flavoprotein, OYEs are used as biocatalysts for decades to reduce activated C=C bonds in α,β-unsaturated carbonyl compounds. The promiscuity of the enzymatic catalysis is advantageous for industrial applications. However, the physiological function of OYEs, especially in bacteria, is still puzzling. Moreover, the relevance of the OYEs in infection conditions remained enigmatic.   Here, I show that there are two groups of OYEs (OYE flavin oxidoreductase, OfrA and OfrB) that are encoded in staphylococci and some firmicutes. OfrA (SAUSA300_0859) is more conserved than OfrB (SAUSA300_0322) in staphylococci and is a part of the staphylococcal core genome. A reporter system was established to report for ofrA in S. aureus background. The results showed that ofrA is induced under electrophilic, oxidative, and hypochlorite stress. OfrA protects S. aureus against quinone, methylglyoxal, hydrogen peroxide, and hypochlorite stress. Additionally, the results provide evidence that OfrA supports thiol-dependent redox homeostasis. At the host-pathogen interface, OfrA promotes S. aureus fitness in murine macrophage cell line. In whole human blood, OfrA is involved in S. aureus survival indicating a potential clinical relevance to bacteraemia. In addition, ofrA mutation affects the production of the virulence factor staphyloxanthin via the upper mevalonate pathway. In summary, decoding OfrA function and its proposed mechanism of action in S. aureus shed the light on a conserved stress response within multiple organisms.}, subject = {Staphylococcus aureus}, language = {en} } @article{IbrahimOhlsen2022, author = {Ibrahim, Eslam S. and Ohlsen, Knut}, title = {The old yellow enzyme OfrA fosters Staphylococcus aureus survival via affecting thiol-dependent redox homeostasis}, series = {Frontiers in Microbiology}, volume = {13}, journal = {Frontiers in Microbiology}, issn = {1664-302X}, doi = {10.3389/fmicb.2022.888140}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-274381}, year = {2022}, abstract = {Old yellow enzymes (OYEs) are widely found in the bacterial, fungal, and plant kingdoms but absent in humans and have been used as biocatalysts for decades. However, OYEs' physiological function in bacterial stress response and infection situations remained enigmatic. As a pathogen, the Gram-positive bacterium Staphylococcus aureus adapts to numerous stress conditions during pathogenesis. Here, we show that in S. aureus genome, two paralogous genes (ofrA and ofrB) encode for two OYEs. We conducted a bioinformatic analysis and found that ofrA is conserved among all publicly available representative staphylococcal genomes and some Firmicutes. Expression of ofrA is induced by electrophilic, oxidative, and hypochlorite stress in S. aureus. Furthermore, ofrA contributes to S. aureus survival against reactive electrophilic, oxygen, and chlorine species (RES, ROS, and RCS) via thiol-dependent redox homeostasis. At the host-pathogen interface, S. aureusΔofrA has defective survival in macrophages and whole human blood and decreased staphyloxanthin production. Overall, our results shed the light onto a novel stress response strategy in the important human pathogen S. aureus.}, language = {en} }