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2D-LC/MS/MS analysis of Escherichia coli during exposure an abrupt downshift in water activity

ABSTRACT: 2D-LC/MS/MS analysis was used to examine time-dependent changes in proteome of E. coli O157:H7 strain Sakai upon an abrupt downshift in water activity (aw) (i.e., from aw 0.993 to aw 0.967) at a constant temperature of 35°C. Bacterial cells were harvested before hyperosmotic shift, and 0 (i.e. immediately after the shift), 30, 80, and 310 min after the shift. It also should be noted that these time points were chosen with the aims to characterize the physiology of E. coli during dynamic changes in growth kinetics induced by an abrupt downshift in water activity. Specifically, the samples taken at time 0 and 30 min were obtained during the period in which the loss of cell culturability was observed, whereas the samples at time 80 and 310 min respectively reflected the physiological state of E. coli during the ‘recovery’ period and during growth after the shift.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Escherichia Coli

SUBMITTER: Chawalit Kocharunchitt

PROVIDER: PXD000541 | Pride | 2014-04-14

REPOSITORIES: Pride

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Global genome response of Escherichia coli O157∶H7 Sakai during dynamic changes in growth kinetics induced by an abrupt downshift in water activity.

Kocharunchitt Chawalit C King Thea T Gobius Kari K Bowman John P JP Ross Tom T

PloS one 20140303 3

The present study was undertaken to investigate growth kinetics and time-dependent change in global expression of Escherichia coli O157∶H7 Sakai upon an abrupt downshift in water activity (aw). Based on viable count data, shifting E. coli from aw 0.993 to aw 0.985 or less caused an apparent loss, then recovery, of culturability. Exponential growth then resumed at a rate characteristic for the aw imposed. To understand the responses of this pathogen to abrupt osmotic stress, we employed an integr ...[more]

PMID: 24594867

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2D-LC/MS/MS analysis of Escherichia coli during exposure to an abrupt temperature downshift

Project description:2D-LC/MS/MS analysis was used to examine time-dependent changes in proteome of E. coli O157:H7 strain Sakai upon an abrupt downshift in temperature (i.e., from 35°C to 14°C). Cell cultures were harvested at 30, 90, 160, and 330 min post-temperature downshift. It also should be noted that these time points were chosen with the aims to characterize the physiology of E. coli during dynamic changes in growth kinetics induced by an abrupt temperature downshift. Specifically, the samples taken at time 30 and 90 min were obtained during adaptation period, whereas the samples at time 160 and 330 min reflected the physiological state of E. coli during growth after the shift. MS/MS data obtained from each protein sample were processed by the Computational Proteomics Analysis System (CPAS), a web-based system built on the LabKey Server (v9.1, released 02.04.2009). The experimental mass spectra produced were subjected to a semi-tryptic search against the combined databases of E. coli O157:H7 Sakai (5,318 entries in total) downloaded from the National Center for Biotechnology Information (NCBI, https://www.ncbi.nlm.nih.gov/, downloaded 25.11.2008) using X!Tandem v2007.07.01. These databases included the E. coli O157:H7 Sakai database (5230 entries, NC_002695.fasta) and two E. coli O157:H7 Sakai plasmid databases, plasmid pO157 (85 entries, NC_002128.fasta) and plasmid pOSAK1 (three entries, NC_002127.fasta). The parameters for the database search were as follows: mass tolerance for precursor and fragment ions: 10 ppm and 0.5 Da, respectively; fixed modification: cysteine cabamidomethylation (+57 Da); and no variable modifications. The search results were then analyzed using the PeptideProphet and ProteinProphet algorithms from the Trans Proteomic Pipeline v3.4.2. All peptide and protein identifications were accepted at PeptideProphet and ProteinProphet of ≥0.9, corresponding to a theoretical error rate of ≤2%.

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2D-LC/MS/MS analysis of Escherichia coli during exposure to abrupt downshifts in temperature and water activity

Project description:2D-LC/MS/MS analysis was used to examine time-dependent changes in proteome of E. coli O157:H7 strain Sakai upon abrupt downshifts from 35°C aw 0.993 to 14°C aw 0.967. Bacterial cells were harvested before abrupt downshifts in both temperature and aw (i.e. control), or at 0 (i.e. immediately after the shift), 60, 250, 1605, 4,070, 5,700, 9,900 or 18,565 min after the shifts.

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Global genome response of Escherichia coli to water activity downshift

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