Mutations in atpG Affect Postranscriptional Expression of pckA in Escherichia coli
Spontaneous mutants isolated with Pck- (lack of PEP carboxykinase activity) and Suc- (inability to utilise succinate as carbon source) phenotypes were previously characterised as atpG mutants defective in the ã subunit of ATP synthase.
In this work we find by reverse transcriptase and real time quantitative PCR that levels of pckA mRNA are normal in the atpG mutants and that the defects in expression of pckA are therefore likely at the level of translation, protein assembly and/or protein degradation. As expected, ATP synthase activity and proton pumping in inside-out membrane vesicles were defective in these atpG mutants. It is likely that one of these defects is affecting regulation or expression of the pckA gene. It was observed that atpG mutants were defective in calcium-dependent transformation although they could be made competent for electroporation. The atpG mutants were also defective for growth of P1 bacteriophage although they could serve as recipients for P1-dependent generalised transduction. These latter phenotypes are also likely due to defects in energy metabolism.
Advisor:Dmitriev, Oleg; Howard, Peter; Bull, Harold; Goldie, Hughes
School:University of Saskatchewan
School Location:Canada - Saskatchewan
Source Type:Master's Thesis
Keywords:fluorescence quenching real time reverse transcriptase pcr gene regulation pep carboxykinase atpg mutants atp synthase gluconeogenesis escherichia coli genetics acridine orange
Date of Publication:05/05/2008