Comparative proteomic analysis of Bacillus thuringiensis wild-type and two mutant strains disturbed in polyphosphate homeostasis

Authors: FİLİZ YEŞİLIRMAK, TUĞRUL DORUK, ŞERİF YILMAZ, SEDEF TUNCA GEDİK

Abstract: Polyphosphate polymer (polyP) plays a very important role in every living cell. Synthesis of this linear polymer of phosphate (P$_{i}$) residues is catalyzed by the polyphosphate kinase (PPK) enzyme. It was shown that high levels of intracellular polyphosphate stimulated endotoxin production by Bacillus thuringiensis subsp. israelensis (Bti). In this study, proteomic analysis of the wild-type and two mutant strains, overexpressing the ppk gene (Bti pHTppk) and without the ppk gene (Bti Δppk), were used to clarify the relation between polyP and endotoxin production. Intracellular proteins were separated by two-dimensional gel electrophoresis; 41 spots of interest (proteins differentially expressed) were obtained and 35 of them were identified by mass spectrometry. Analysis of the protein profiles showed that there is a general decrease in the expression levels of proteins related with energy metabolism, amino acid metabolism, and purine biosynthesis in both Bti pHTppk and Bti Δppk. Gluconeogenesis and fatty acid metabolism were also slowed down in both strains, whereas expression of stress response proteins increased compared to the wild-type. These results suggested that changes in polyP concentration cause a general stress condition inside the cell, which in turn stimulates secondary metabolite synthesis.

Keywords: Polyphosphate polymer, polyphosphate kinase, two-dimensional gel electrophoresis, endotoxin, bioinsecticide, Bacillus thuringiensis israelensis, secondary metabolite, regulation, proteomics

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