Konstrukce neoznačených mutantů v genech kódujících Ser/Thr proteinkinasy a fosfoproteinfosfatasu Pseudomonas aeruginosa a analýza jejich vlastností

Key Points

Protein phosphatase gene stp1 and ser/thr protein kinases stk1, ppkB were targeted by gene replacement techniques.
Single mutants were successfully created using Flp-FRT recombination, allowing for subsequent antibiotic cassette excision.
Two-dimensional gel electrophoresis enabled the analysis of phosphoprotein patterns from both wild type and mutant strains.
These findings highlight the critical role of these enzymes in the physiology of Pseudomonas aeruginosa.

Abstract

To investigate the importance of individual protein kinases and phosphatases in P. aeruginosa physiology we prepared the single and double mutants in the coresponding genes. Single mutants in protein kinase genes stk1, ppkB and protein phosphatase gene stp1 were prepared by the method of gene replacement with subsequent excision of antibiotic cassettes using Flp-FRT recombination system. Two-dimensional gel electrophoresis was used to analyze the patterns of phosphoproteins obtained in in vitro protein kinase assays with the cell-free lysates of wild type as well as all mutant strains.

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Konstrukce neoznačených mutantů v genech kódujících Ser/Thr proteinkinasy a fosfoproteinfosfatasu Pseudomonas aeruginosa a analýza jejich vlastností

Key Points

Abstract

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