Comparative proteomics of oxidative stress response of Lactobacillus acidophilus NCFM reveals effects on DNA repair and cysteine de novo synthesis

Abstract

Probiotic cultures encounter oxidative conditions during manufacturing, yet protein abundance changes induced by such stress have not been characterized for some of the most common probiotics and starters. This comparative proteomics investigation focuses on the response by Lactobacillus acidophilus NCFM to H O simulating an oxidative environment. Bacterial growth was monitored by BioScreen and batch cultures were harvested at exponential phase for protein profiling of stress responses by 2D gel based comparative proteomics. Proteins identified in 19 of 21 spots changing in abundance due to H O were typically related to carbohydrate and energy metabolism, cysteine biosynthesis, and stress. In particular, increased cysteine synthase activity may accumulate a cysteine pool relevant for protein stability, enzyme catalysis, and the disulfide-reducing pathway. The stress response further included elevated abundance of biomolecules reducing damage such as enzymes from DNA repair pathways and metabolic enzymes with active site cysteine residues. By contrast, a protein-refolding chaperone showed reduced abundance, possibly reflecting severe oxidative protein destruction that was not overcome by refolding. The proteome analysis provides novel insight into resistance mechanisms in lactic acid bacteria against reactive oxygen species and constitutes a valuable starting point for improving industrial processes, food design, or strain engineering preserving microorganism viability.

Complex Responses to Hydrogen Peroxide and Hypochlorous Acid by the Probiotic Bacterium Lactobacillus reuteri

“…L. reuteri’s response to H 2 O 2 was generally consistent with what has been previously observed with other catalase-negative Gram-positive bacteria (21, 23, 24, 31, 65, 67, 68). Highly upregulated genes included genes encoding alkylhydroperoxidase ( ahpCF ) (69), NADH oxidase ( noxE ) (70), and methionine sulfoxide reductase ( msrB ) (47), DNA repair genes ( uvrABD , xthA , and umuC ) (71), and genes for predicted metal transporters ( pcl1 and pcl2 ) and the peroxide-sensing transcription factor PerR (65).…”

“…This is especially true of RCS, including hypochlorous acid (HOCl) and reactive chloramines, which are extremely potent antimicrobial compounds produced by the neutrophil enzyme myeloperoxidase (22, 26–28). Relatively little is known about how health-associated probiotic and commensal bacteria sense and respond to inflammatory oxidants (21, 29–31).…”

Probiotics alter biofilm formation and the transcription of Porphyromonas gingivalis virulence-associated genes

“…Negative regulation of kgp and rgpA transcription in W83 was achieved in most conditions by L. acidophilus LA5, whereas other probiotics even up-regulated transcription of these proteases encoding-genes. In fact, a proteomics of L. acidophilus revealed that an increased cysteine synthase activity may accumulate a cysteine pool relevant for protein stability and enzyme catalysis in this probiotic, which suggests its advantage in terms of protease regulation [ 50 ]. Still, the decreased expression of kgp and rgp induced by some probiotics, especially L. acidophilus LA5, may not only impair uptake of iron and decrease tissue destruction by the periodontopathogen strains but should also play an important role adherence of P. gingivalis to epithelial cells [ 20 , 51 ], in consonance with our previous data [ 9 ].…”

Intra-species Genomic and Physiological Variability Impact Stress Resistance in Strains of Probiotic Potential

“…Universal bacterial stress-response systems encode mechanisms to cope with the deleterious consequences of exposure to environmental stresses, including DNA damage, protein misfolding, and loss of cell wall/membrane integrity. Bacterial DNA damage is repaired by MutS and RecA-like recombinases (Lee and Pi, 2010 ; Rossi et al, 2016 ; Calderini et al, 2017 ; Overbeck et al, 2017 ). Protein misfolding is reduced and repaired by molecular chaperones including heat-shock proteins (Ruiz et al, 2013 ; Calderini et al, 2017 ).…”

“…Bacterial DNA damage is repaired by MutS and RecA-like recombinases (Lee and Pi, 2010 ; Rossi et al, 2016 ; Calderini et al, 2017 ; Overbeck et al, 2017 ). Protein misfolding is reduced and repaired by molecular chaperones including heat-shock proteins (Ruiz et al, 2013 ; Calderini et al, 2017 ). Cell wall integrity of gram positive bacteria is maintained through lipoteichoic acid (LTA), exopolysaccharide, and fatty acid metabolism/synthesis genes (Koskenniemi et al, 2011 ).…”