BayBioMS - Bacteriocin peptides Lactobacillus curvatus

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Bacteriozin_peptides_Lactobacillus_curvatus_P099_01_2020-01-15_16-17-28.sky.zip2020-02-06 09:30:48488241
Interspecies assertiveness of Lactobacillus curvatus and Lactobacillus sakei in sausage fermentations

  • Organism: Lactobacillus curvatus subsp. curvatus
  • Instrument: Q Exactive HF-X
  • SpikeIn: No
  • Keywords: Lactobacillus curvatus, Lactobacillus sakei, fermented sausage, assertiveness, stress tolerance, bacteriocin
  • Lab head: Christina Ludwig
Abstract
Lactobacillus (L.) curvatus and L. sakei contain strains, which are assertive in sausage fermentation. Previous work has demonstrated differences in assertiveness at strain level within one species, and revealed either exclusion of competitors by complementary partner strains or their inhibition by single strains. This work addresses interspecies differences in the assertiveness of L. curvatus and L. sakei. Strain sets of L. curvatus and L. sakei were employed as starters in a fermented sausage model and their abundancy upon fermentation was determined by strain-specific MALDI-TOF MS identification. Generally, single or groups of L. sakei strains outcompeted L. curvatus strains. In multiple growth tests employing mMRS and mMSM it could be shown that assertive L. sakei strains can be predicted along their µ max in mMSM. Still, L. curvatus TMW 1.624 could suppress all L. curvatus and most L. sakei strains in competitive settings. This could be referred to its expression of several bacteriocins, which are active against all of the L. curvatus strains. Strain specific differences could be demonstrated in the susceptibility of L. sakei to bacteriocins, and in oxidative stress tolerance, which is higher in co-existing L. sakei strains than in the bacteriocin producer. This suggests that tolerance to bacteriocins and oxidative stress represent additional determinants for assertiveness, above previously reported bacteriocin production versus metabolic complementarism of partner strains.
Experiment Description
The expression of different bacteriocins and their extracellular release into the growth medium was investigated by label free quantitative proteomics, following the protocol of Heinze et al. (Heinze et al., 2018). Briefly, L. curvatus TMW 1.624 was grown in mMSM medium. 2 ml of the culture were harvested by centrifugation (21,100 g, for 5 min at 4 °C), the supernatant was collected without disrupting the cell pellet and mixed with LDS sample buffer (Thermo Fisher Scientific) for in-gel digestion. As a negative control pure mMSM medium was analysed. The samples were then reduced, heated, alkylated, and run on a NuPAGE gel. In-gel tryptic digestion, which was performed according to the standard procedures (Shevchenko et al. 2006) and analysed via LC-MS/MS on an Ultimate 3000 RSLCnano system coupled online to a Q-Exactive HFX mass spectrometer (ThermoFisher Scientific) in data-dependent acquisition (DDA) mode.
Sample Description
Peptide identification and quantification were performed using the MaxQuant software (version 1.6.3.4) by searching the MS data against the L. curvatus TMW 1.624 proteome database, including bacteriocin peptide sequences predicted from the genome of L. curvatus TMW 1.624. Confident identification of all 8 Bacteriocin-related peptides was manually validated using the Skyline software. The resulting Skyline document, entailing spectrum information, mass accuracy information and chromatogram information, can be accessed via Panorama Public.
Created on 2/6/20, 9:30 AM

The DDA results associated with this experiments are available in the ProteomeXchanage dataset PXD016990