Intracellular Poly-3-Hydroxybutyrate Played An ImportantRole In Growth Characteristics And Stress Resistances Of Bacillus Thuringiensis Vegetative Cells

The research use Bacillus thuringiensis (Bt) strain BMB171,poly-3-hydroxybutyrate(Poly-3-hydroxybutyrate, PHB) synthetic gene cluster (phaRBC) strengthen strainBMB171+p and PHB synthase gene (phaC) blocked mutant BMB171-p for objects of thisresearch, to investigate the influence of PHB on resistance to the environmental stress ofBt vegetative cells for the purpose, System compared the differences of reverseenvironmental tolerances (including heat, cold-resistant, UV-resistant and hunger) andthe growth capabilities in the different pH medium, osmotic pressure medium, differentconcentrations of organic solvent medium systematicly,, besides analyzing thermaldynamics by differences of thermal tolerances of the three strains, BMB171+p,BMB171and BMB171-p. This is the first systematic study of physiology function of PHB onBacillus thuringiensis vegetative cell stress tolerance and growth abilities in adverseenvironments, and elucidate thermal dynamics of the three kind strains stressingengineering centered, It is the basis to further study the biological function of PHB.Study on the relationship of process of cell growth curve and the fermentation yieldPHB with time curve, the results showed (1) PHB was the primary metabolite of Bt. (2)Strains BMB171 and BMB171+p can synthesize PHB normally in life cycle, at the end oflogarithmic growth phase PHB synthesis achieved the maximum, the maximum were0.844 mg/mL andl.248 mg/mL, respectively, the latter biggest production increase ofnearly 50%; the mutant strain BMB171-p can not synthesize PHB. (3) PHB metabolismaffect organic and carbon source metabolism.(4) BMB171+p and BMB171 strains canform spores normally, but BMB171-p can not and cell autolysis.Study on the stress tolerance of the three kinds vegetative cells, first investigatephysiology function of PHB in adverse environment, the results suggested (1) comparisonof D₉₀ values of strains BMB171+p,BMB171 and BMB171-p, the heat resistance wasBMB171+p＞BMB171＞＞BMB171-p; (2) put three strains vegetative cells at -20℃for five days, the survival rate of strain BMB171+p vegetative cells were 15.91%; strainBMB171 vegetative cells were 14.93%; but strain BMB171-p vegetative cells were only0.93%; (3) the abilities of anti-ultraviolet of BMB171+p and BMB171 were higher thanBMB171-p; (4) BMB171+p and BMB171 had bigger stress tolerance to hunger, butBMB171-p had cell autolysis.This study first investigate the growth curve of BMB171,BMB171+p and BMB171-pstrains by inoculated in the different pH mediums, osmotic pressure mediums, differentconcentrations of organic solvent mediums, discuss influence of growth capabilities of thethree kind strains related to PHB. The experimental results indicate that: the growth ability of each strain was decreased with the degree of adverse conditions, the degree ofBMB171+p and BMB171 strains reduced almost, but the degree of BMB171-p strainreduced much more than others, it proved that the strains produced PHB were more adaptto the environmental changes than that no PHB strains.Select one of the stress tolerance—heat resistance, this research first elucidatethermal dynamics of the three kind strains stressing engineering centered. By analyzingthe heat death rate constant k values and the activate energyâ–³E of BMB171+p,BMB171and BMB171-p strains, the results show: at the same temperature, k_BMB171+p＜k_BMB171＜＜k_BMB171-p,â–³E values were 140.3, 131.1 and 108.3 kJ/mol respectively.