Significance of Four Methionine Sulfoxide Reductases in Staphylococcus aureus

Abstract

Staphylococcus aureus is a major human pathogen and emergence of antibiotic resistance in clinical staphylococcal isolates raises concerns about our ability to control these infections. Cell wall-active antibiotics cause elevated synthesis of methionine sulfoxide reductases (Msrs: MsrA1 and MsrB) in S. aureus. MsrA and MsrB enzymes reduce S-epimers and R-epimers of methionine sulfoxide, respectively, that are generated under oxidative stress. In the S. aureus chromosome, there are three msrA genes (msrA1, msrA2 and msrA3) and one msrB gene. To understand the precise physiological roles of Msr proteins in S. aureus, mutations in msrA1, msrA2 and msrA3 and msrB genes were created by site-directed mutagenesis. These mutants were combined to create a triple msrA (msrA1, msrA2 and msrA3) and a quadruple msrAB (msrA1, msrA2, msrA3, msrB) mutant. These mutants were used to determine the roles of Msr proteins in staphylococcal growth, antibiotic resistance, adherence to human lung epithelial cells, pigment production, and survival in mice relative to the wild-type strains. MsrA1-deficient strains were sensitive to oxidative stress conditions, less pigmented and less adherent to human lung epithelial cells, and showed reduced survival in mouse tissues. In contrast, MsrB-deficient strains were resistant to oxidants and were highly pigmented. Lack of MsrA2 and MsrA3 caused no apparent growth defect in S. aureus. In complementation experiments with the triple and quadruple mutants, it was MsrA1 and not MsrB that was determined to be critical for adherence and phagocytic resistance of S. aureus. Overall, the data suggests that MsrA1 may be an important virulence factor and MsrB probably plays a balancing act to counter the effect of MsrA1 in S. aureus.

Biotin sulfoxide reductase contributes to oxidative stress tolerance and virulence in Salmonella enterica serovar Typhimurium

“…This might be explained by the high concentrations of H 2 O 2 that we used for the survival assays in vitro. Other studies addressing the role of bacterial methioinine sulfoxide reductases in the oxidative stress response use similar or even higher concentrations of H 2 O 2 , which are presumably required because of the redundancy of methionine sulfoxide repairing enzymes (Denkel et al, 2011;Douglas et al, 2004;Lee et al, 2009;Singh & Singh, 2012;Zhao et al, 2010). Methionine is susceptible to oxidation in its free and protein-bound form (Weissbach et al, 2002).…”

Effect of clpP and clpC deletion on persister cell number in Staphylococcus aureus

“…at 37 C for the duration of all growth experiments (Controlled Environment Incubator Shaker; New Brunswick Scientific). All samples from various S. aureus cultures were diluted by serial 10-fold dilutions and plated on Singh & Singh (2012) *S. aureus strains were suspended in TSB containing 10 % glycerol for long-term storage and kept at À70 C. Frozen cultures were thawed at room temperature. Fifty microlitres of each S. aureus strain was inoculated in 5 ml of TSB and incubated overnight (16-18 h) at 37 C.…”

“…The apparent lack of persister cells in early exponential phase may be due, in part, to low concentrations of persister proteins that may act as quorum-sensing molecules (Lewis, 2007). Others have shown that two processes may influence persister formation: a stochastic variation in the level of specific persister proteins and a controlled, regulated threshold level of these proteins which may be dependent on population density (Lewis, 2007;Tashiro et al, 2012).…”

Staphylococcus aureus SaeR/S-regulated factors reduce human neutrophil reactive oxygen species production

“…The importance of detoxifying ROS is demonstrated by the many mechanisms used by S. aureus that include scavenging and neutralizing ROS, such as O 2 2 and H 2 O 2 , with SODs (SodA and SodM) and catalase (KatA), respectively [37,38]. In addition, the iron-regulated surface-determinant proteins IsdA and IsdB have been implicated in increasing S. aureus resistance to killing by H 2 O 2 [21], as well as methionine sulfoxide reductases to reduce oxidized methionine residues following oxidative stress that increase S. aureus resistance to ROS [39]. Unlike previously described mechanisms to neutralize ROS, our data suggest that S. aureus can directly inhibit H 2 O 2 and HOCl production, resulting in increased S. aureus survival.…”