Purification and characterization of the Staphylococcus aureus bacillithiol transferase BstA

Area of Science:

• Microbiology
• Biochemistry
• Enzymology

Background:

• Firmicutes bacteria, including Staphylococcus aureus, utilize bacillithiol (BSH) as their primary low molecular weight thiol.
• The bacillithiol transferase YfiT from Bacillus subtilis belongs to the recently identified DinB/YfiT-like Superfamily.
• Most Staphylococcus aureus strains possess a gene encoding a bacillithiol transferase within the DinB/YfiT-like Superfamily.

Purpose of the Study:

• To characterize the bacillithiol transferase (BstA) from Staphylococcus aureus Newman.
• To investigate the enzymatic activity, substrate specificity, and kinetic properties of BstA.
• To explore the potential role of BstA in bacterial detoxification pathways.

Main Methods:

• Cloning, purification, and activity confirmation of recombinant BstA.
• In vitro biochemical assays to study saturation kinetics and substrate specificity.
• Enzyme activity measurements in Staphylococcus aureus cells under different growth conditions.

Main Results:

• Recombinant BstA demonstrated activity with bacillithiol but not other tested thiols.
• BstA catalyzed the conjugation of bacillithiol to model substrates like monochlorobimane and dinitrobenzene, and the antibiotic cerulenin.
• Bacillithiol alone could react with some electrophilic molecules, but BstA activity was low in nutrient-rich media.

Conclusions:

• BstA is an active bacillithiol transferase from Staphylococcus aureus Newman, capable of detoxifying cerulenin.
• Bacillithiol itself may directly detoxify certain electrophilic compounds.
• BstA represents the first identified member of the DinB/YfiT-like Superfamily from Staphylococcus aureus and is distinct from B. subtilis YfiT.