Dataset Information

Pulsed SILAC proteomics analysis of Escherichia coli RPL11 methyltransferase PrmA mutant

ABSTRACT: For the pulse-chase experiment, E. coli BW25113 and its isogenic delta prmA strains were grown in M9 minimal medium and L-Lysine-2HCl (13C6, 15N2) was added to cultures in exponential and stationary phase. The cells were cultured for an additional 2 hr and harvested. Proteins from the wild-type and delta prmA strains were extracted, digested with trypsin, and analyzed by LC-MS/MS on a Q-Exactive HF-X. LC-MS/MS data was processed with MaxQuant v2.2.0.0, identifying peptides by searching tandem mass spectra against the E. coli K12 sequences from Uniprot Knowledgebase downloaded on December 19, 2022. The search parameters included fully LysC digestion with 2 missed cleavage sites, oxidation of methionine as variable modifications, and precursor mass tolerances of 20 ppm and 4.5 ppm for the first and main searches, respectively. Multiplicity was set as 2 of a light channel, and a heavy channel with Lys8. The "requantify" function was enabled.

INSTRUMENT(S): Q Exactive HF-X

ORGANISM(S): Escherichia Coli Bw25113 (ncbitaxon:679895)

SUBMITTER: Ernesto Nakayasu

PROVIDER: MSV000094475 | MassIVE | Fri Apr 05 14:37:00 BST 2024

REPOSITORIES: MassIVE

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Project description:Leafy green vegetables, such as lettuce, have been increasingly implicated in outbreaks of foodborne illnesses due to contamination by Escherichia coli O157:H7. While E. coli can survive in soils, colonize plants, and survive on produce, very little is known about the interaction of E. coli with the roots of growing lettuce plants. In these studies, a combination of microarray analyses and surface enhanced Raman spectroscopy (SERS) were used to gain a comprehensive understanding of bacterial genes involved in the colonization and growth of E. coli O157:H7 on lettuce roots and compared to E. coli K12 using a hydroponic system (HS) which we have reported in the previous studies. Using microarray, after three days of interaction with lettuce roots, 94 and 109 genes of E. coli O157:H7 were significantly up-regulated and down-regulated at least 1.5 fold, respectively. Only 8 genes were also found in the E. coli K12 up-regulated genes. No genes were found in the down-regulated genes clusters between those two strains. For E. coli O157:H7, forty out of the 94 up-regulated genes (43%) were involved in protein synthesis and were highly repressed compared to 40 out of 193 (23%) E. coli K12 up-regulated genes associated with protein synthesis. The wildtype of E.coli O157:H7 colonized two log CFU per root less compared to E. coli K12. Genes involved in biofilm modulation (bhsA and ybiM) were significantly up-regulated in E. coli O157:H7 and curli production (crl and csgA) were found important for E. coli K12 to attach to lettuce roots in the previous studies. BhsA mutant of E. coli O157:H7 was impaired in the colonization of lettuce roots. The SERS spectra of E. coli K12 and O157 controls (cells without interacting with roots) were very similar. The spectra of E. coli K12 and O157 exposed to the hydroponic system (HS) showed some differences in the nucleic acid, protein, and lipid regions compared with controls. The spectra of E. coli K12 HS cells exhibited significant differences compared to spectra from E. coli O157 HS cells in the RNA and protein regions. The overall band intensity of amide regions declined for E. coli O157 HS cells, while it increased for E. coli K12 HS cells. The intensity of the RNA bands of E. coli K12 HS cells were also found much higher than those of E. coli O157 HS cells. These findings were in agreement to our Microarray data. Our microarray and SERS data showed that E. coli K12 and O157:H7 behavior dramatically differently in colonizing on lettuce roots. Compared to K12, E. coli O157:H7 colonized less efficiently on lettuce roots.

Dataset Information

Pulsed SILAC proteomics analysis of Escherichia coli RPL11 methyltransferase PrmA mutant

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets