PHENOTYPIC ANALYSIS OF CLPX, SIG E AND SIG B MUTANTS OF PROPIONIBACTERIUM FREUDENRICHII
2025
Rajapaksha, Supun | Helsingin yliopisto, Maatalous-metsätieteellinen tiedekunta | University of Helsinki, Faculty of Agriculture and Forestry | Helsingfors universitet, Agrikultur-forstvetenskapliga fakulteten
Propionibacterium freudenreichii, a bacterium with significant applications in food biotechnology and as a probiotic, encounters a range of environmental stresses throughout its life cycle. While understanding its stress response is critical, the roles of key regulatory elements, specifically the sigma factors; SigB and SigE, and the ClpX ATPase involved in protein quality control, remain poorly defined in this species. This study aimed to characterize the growth and stress resistance of ΔclpX, ΔsigE, and ΔsigB mutants in P. freudenreichii DSM4902 to elucidate their potential regulatory roles in key phenotypes. Growth kinetics and quantitative assessment of viable cell counts via spot dilution assays were employed to compare the growth and viability of wild-type and mutant strains under various stress conditions, including thermal (heat and cold), osmotic, bile, acidic pH, puromycin, ethanol and oxidative stress. Statistical analysis of final biomass data was performed using the Mann-Whitney U test. The results revealed that both SigB and SigE had no effect towards the bile stress adaptation while contributed to the puromycin stress tolerance. SigE had no involvement in aerobic stress while SigB was inactive towards the anaerobic conditions. Notably, ClpX broadly influenced general stress tolerance except for puromycin and aerobic stress. All three regulons were essential for high temperature stress adaptation even evident with a statistical significance (p ≤ 0.05). Role of ClpX, SigE, and SigB, found to be important in handling low pH (pH 5.2) and 3% ethanol stress. These findings highlight the distinct and overlapping roles of ClpX, SigE, and SigB in the stress response of P. freudenreichii.
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