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Proteomics analysis of Escherichia coli RPL11 methyltransferase PrmA mutant

ABSTRACT: E. coli BW25113 and its isogenic delta prmA strains were grown in 5 mL TB7 (10 g/L tryptone buffered at pH 7.0 with 100 mM potassium phosphate) at 37 deg C, aerated at 225 rpm for 16 h. Cells were harvested by centrifugation at 9,400 x g, washed once with PBS and stored at -80 deg C for proteomics analysis. E. coli proteins from the wild-type and delta prmA strains were extracted, digested with trypsin, and analyzed by LC-MS/MS on a Q-Exactive Plus, as described in detail by Yang et al. (doi: 10.1111/1462-2920.13852). 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 tryptic digestion with 2 missed cleavage sites, oxidation of methionine, and lysine tri-methylation as variable modifications, and precursor mass tolerances of 20 ppm and 4.5 ppm for the first and main searches, respectively. Label-free quantification was set as default parameters and the "matching between runs" function was enabled using a matching window of 3 min and an alignment window of 20 min.

INSTRUMENT(S): Q Exactive Plus

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

SUBMITTER: Ernesto Nakayasu

PROVIDER: MSV000092270 | MassIVE | Mon Jun 26 14:21:00 BST 2023

REPOSITORIES: MassIVE

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Proteomics analysis of Escherichia coli RPL11 methyltransferase PrmA mutant

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