Abstract
Most bacterial cells are 1-2 microns in size, limiting intracellular products like polyhydroxyalkanoates (PHA) accumulation. Cell size is regulated by key genes such as mreB and minCD, which encode cellular skeleton protein and control cell fission ring location, respectively. Their expression changes significantly affect microbial growth. This study successfully redesigns the ClpXP protein degradation system by deleting the sspB gene and using mutated SsrA tags with different degradation rates to control MreB degradation. Dynamic degradation of MreB allows non-model bacterium Halomonas bluephagenesis to grow normally and increase cell size simultaneously. Combined with overexpression of minCD, H. bluephagenesis with progressive MreB degradation increases the cell size further, albeit with a reduced growth rate. H. bluephagenesis CYL0307, with the PHB granule-associated protein PhaP1 deleted and phaAB(Re) overexpressed in the MreB-degraded strain, increases cell volume more than nine times compared to the original strain. CYL0307 produces 149 g L(-1) cell dry weight containing 82% PHB after 44 h in a 5000 L bioreactor, with cells containing single large PHB granules, simplifying recovery and purification. These results provide a post-translational gene regulation method in H. bluephagenesis and a strategy for enhancing PHB production via morphological engineering.