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Nonlabeled Quantitative Proteomic Comparison Identifies Differences in Acid Resistance Between Escherichia coli O157:H7 Curli Production Variants
- Gunther Nereus W., Liu Yanhong, Nunez Alberto, Paul Moushumi, Uhlich Gaylen A.
- Foodborne pathogens & disease 2014 v.11 no.1 pp. 30-37
- Escherichia coli O157, acid tolerance, bacteria, bacterial proteins, fractionation, gene expression regulation, genes, knockout mutants, pH, phenotype, protein synthesis, proteomics, spectrometers, transcription (genetics)
- To understand the nature of a bacterial strain, it is necessary to be able to identify and measure the proteins expressed by the bacteria. In this research, the entire protein complements produced by Escherichia coli O157:H7 strain 43894OW and its naturally occurring curli producing variant 43894OR were compared to better understand the unique capabilities of these two closely related strains. A nonlabeled proteomic comparison was performed utilizing the spectra counting and peptide fractionation abilities of a quadrupole-time of flight analyzer mass spectrometer to identify and quantitate the proteins produced by the two strains. The process reliably identified and measured the concentration of 419 proteins from strains 43894OW and 43894OR within three separate biological replicates. From these two sets, 59 proteins were identified that were preferentially expressed in strain 43894OW compared to 43894OR and 14 proteins that were conversely preferentially expressed in 43894OR. A subset of the preferentially expressed proteins was assayed to determine whether their levels of gene transcription corresponded with the observed protein expression. From the resulting list of confirmed differentially expressed proteins, it was observed that the proteins contributing to acid survival—GadA and GadB—were overexpressed in 43894OW compared to 43894OR. The predicted enhanced acid resistance phenotype of 43894OW was confirmed by experimentation at pH 2.5. Additionally, a knockout mutation in the csgD genes of the 43894OR strain was constructed and suggested that CsgD had a repressive effect on acid survival in 43894OR.