Studies of Universal Stress Protein and Poly-3-hydroxybutyrate in Bacillus thuringiensis strains
Bacillus thuringiensis (Bt) produces the Universal stress protein (USP) in response to growth inhibition and poly-3-hydroxybutyrate (PHB) to provide energy through the process of sporulation. It was hypothesized that the presence of USP positively effects the production of PHB. The main objective of the work reported here was to determine if a correlation exists between USP and PHB production in Bt by detection of these molecules. The presence of PHB in six Bt strains (ATCC 19266, 19267, 19268, 19270, 33679, and 35646) was verified by growth in PHB production medium and heat-fixed slides stained with Nile Blue A viewed by fluorescence microscopy. E. coli (ATCC 35646) served as a negative control. It was noted that intracellular fluorescent particles existed in the cells of all of the Bt strains tested. Proteomic experiments included the detection of USP and PhaC, the PHB synthase. A Sigma Aldrich CelLytic B Plus kit was used to extract bacterial protein and silver staining SDS PAGE gels verified the presence of proteins. Custom polyclonal antibodies produced by GenScript were used for immunodetection of target proteins via Western blot. It was noted that both USP and PhaC were detected in all Bt strains grown in the PHB production medium for six hours. A sodium hypochlorite-chloroform method was used to extract PHB from bacterial cells along with dry cell weight procedures to obtain quantitative data. It was determined that these Bt strains can produce between 30 and 90% PHB per dry cell weight when grown in PHB production medium for 48 hours. USP was also detected in all strains from these cultures using immunodetection methods. It was also noted that Bt strain ATCC 19267 produced greater amounts of PHB than the other strains tested. It was concluded that a possible correlation exists between Universal stress protein and poly-3-hydroxybutyrate production in strains of Bacillus thuringiensis. Biochemical testing of stress response and fatty acid metabolism should be performed to further investigate this possible relationship.
Letimicia Satiana Fears,
"Studies of Universal Stress Protein and Poly-3-hydroxybutyrate in Bacillus thuringiensis strains"
ETD Collection for Tennessee State University.