Methionine Residues in Exoproteins and Their Recycling by Methionine Sulfoxide Reductase AB Serve as an Antioxidant Strategy in Bacillus cereus

Abstract

During aerobic respiratory growth, Bacillus cereus is exposed to continuously reactive oxidant, produced by partially reduced forms of molecular oxygen, known as reactive oxygen species (ROS). The sulfur-containing amino acid, methionine (Met), is particularly susceptible to ROS. The major oxidation products, methionine sulfoxides, can be readily repaired by methionine sulfoxide reductases, which reduce methionine sulfoxides [Met(O)] back to methionine. Here, we show that methionine sulfoxide reductase AB (MsrAB) regulates the Met(O) content of both the cellular proteome and exoproteome of B. cereus in a growth phase-dependent manner. Disruption of msrAB leads to metabolism changes resulting in enhanced export of Met(O) proteins at the late exponential growth phase and enhanced degradation of exoproteins. This suggests that B. cereus can modulate its capacity and specificity for protein export/secretion through the growth phase-dependent expression of msrAB. Our results also show that cytoplasmic MsrAB recycles Met residues in enterotoxins, which are major virulence factors in B. cereus.

Rhodobacter sphaeroides methionine sulfoxide reductase P reduces R- and S-diastereomers of methionine sulfoxide from a broad-spectrum of protein substrates

“…Briefly, each band was destained with ultra-pure water, reduced with DTT, treated with iodoacetamide, and then proteolyzed with Gold Mass Spectrometry Grade Trypsin (Promega) in the presence of 0.01% ProteaseMAX surfactant (Promega). Peptides were immediately subjected to tandem mass spectrometry as previously recommended to avoid methionine oxidation [36]. The resulting peptide mixtures were analyzed in a data-dependent mode with a Q-Exactive HF tandem mass spectrometer (Thermo) coupled on line to an Ultimate 3000 chromatography system chromatography (Thermo) essentially as previously described [37].…”

The Role of Methionine Sulfoxide Reductases in Oxidative Stress Tolerance and Virulence of Staphylococcus aureus and Other Bacteria

“…Expression of MSR genes in other pathogenic species has also been investigated. Similar to S. aureus MSRA1 / MSRB , in H. pylori [ 20 ], E. coli [ 28 ], and B. subtilis [ 29 ] the late log and stationary phase cultures of these bacteria had the highest MSR activity. In addition to MSRA1 / MSRB induction by cell wall-active antibiotics, piplartine, a biologically active alkaloid from peppers [ 30 ], showed enhanced MSR expression in the pathogenic parasite, Trypanosoma cruzi [ 31 ].…”

Time-course proteomics dataset to monitor protein-bound methionine oxidation in Bacillus cereus ATCC 14579

“…Cell pellets and supernatants were separated by centrifugation (10,000 g , 10 min, 4 °C). Exoproteins were isolated from filtered supernatants by TCA precipitation and soluble cellular proteins were extracted from cell lysates as described previously [1] , [2] , [5] , [6] . To avoid any artefactual oxidation, exoproteins and cellular proteins were resolved on denaturing NuPAGE 4–12% Bis-Tris gels (Invitrogen, ThermoFisher, Courtaboeuf, France) during a 5 min electrophoretic migration using NuPAGE antioxidant added to the NuPAGE running buffer.…”

“…This proteomics dataset comprises the whole set of peptides and Met(O) peptides identified in the cellular proteome ( Supplementary Table 1 ) and exoproteome ( Supplementary Table 2 ) of Bacillus cereus ATCC 14579 ΔpBClin15 (Wild-type, [2] ) and its mutant ΔmsrAB [1] . A total of 8720 and 3417 Met(O)peptides were detected in the cellular proteomes and exoproteomes, respectively.…”