Characterization of the β-Lactam Antibiotic Sensor Domain of the MecR1 Signal Sensor/Transducer Protein from Methicillin-Resistant Staphylococcus aureus

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) has evolved two mechanisms for resistance to beta-lactam antibiotics. One is production of a beta-lactamase, and the other is that of penicillin-binding protein 2a (PBP 2a). The expression of these two proteins is regulated by the bla and mec operons, respectively. BlaR1 and MecR1 are beta-lactam sensor/signal transducer proteins, which experience acylation by beta-lactam antibiotics on the cell surface and transduce the signal into the cytoplasm. The C-terminal surface domain of MecR1 (MecRS) has been cloned, expressed, and purified to homogeneity. This protein has been characterized by documenting that it has a critical and unusual Nzeta-carboxylated lysine at position 394. Furthermore, the kinetics of interactions with beta-lactam antibiotics were evaluated, a process that entails conformational changes for the protein that might be critical for the signal transduction event. Kinetics of acylation of MecRS are suggestive that signal sensing may be the step where the two systems are substantially different from one another.

Site-Saturation Mutagenesis of Position V117 in OXA-1 β-Lactamase: Effect of Side Chain Polarity on Enzyme Carboxylation and Substrate Turnover

“…Interestingly, the rate of the rise in anisotropy at the beginning of the assay (presumably some combination of BOCILLIN FL binding and acylation) is also increased by the presence of bicarbonate. Bicarbonate supplementation also leads to a significant increase in the rate of acylation of the MecR sensor protein from S. aureus (32). In order to further probe the role of polar residues in the active site of the BlaR sensors (which have either an asparagine or a threonine at this position), the anisotropy assay was repeated using OXA-1 V117N and V117T.…”

“…The most likely explanation is that the B. licheniformis protein may have additional active site features that augment the slight carbamate-destabilizing effect of threonine. Cha et al note that different sensor proteins can display vastly different acylation rates, and thus subtle differences in active site residue choice may also simply reflect how different sensor proteins are “tuned” to advantage under particular physiological conditions (32). Ultimately, the more striking deacylation-deficiency displayed by V117N over V117T is probably the reason that the majority of sensor proteins have asparagine at this position.…”

Molecular Basis and Phenotype of Methicillin Resistance in Staphylococcus aureus and Insights into New β-Lactams That Meet the Challenge

“…When the bacterium is exposed to a ␤-lactam, a serine residue in the extracellular sensor domain of BlaR1 and MecR1 undergoes irreversible acylation by the ␤-lactam. This results in a conformational change in the sensor domain that is proposed to transmit the signal (i.e., the presence of a ␤-lactam) to the cytoplasmic domain of BlaR1/MecR1 (23,69,167). The molecular details of the signal transmission events are unknown.…”

Regulation of antibiotic resistance in Staphylococcus aureus

“…Induction by the MecR1-controlled system is much slower than by the BlaR1 system, taking hours instead of minutes (Cha et al, 2007;McKinney et al, 2001;Ryffel et al, 1992). The strong repression of mecA by MecI and slow induction kinetics has severe consequences for MRSA, as induction is sometimes not rapid enough to prevent beta-lactam-induced death.…”