An mRNA Display Approach for Covalent Targeting of a Staphylococcus aureus Virulence Factor

Area of Science:

• Biochemistry
• Microbiology
• Medicinal Chemistry

Background:

• Staphylococcus aureus (S. aureus) is a major human pathogen causing over a million deaths annually.
• Fluorophosphonate binding hydrolases (Fphs) are key to S. aureus lipid metabolism and host colonization.
• Fphs are promising targets for novel diagnostics and therapeutics due to their specific expression in Staphylococcus.

Purpose of the Study:

• To develop and screen cyclic peptide libraries for inhibitors of Fphs using mRNA display.
• To create selective covalent inhibitors and fluorescent probes targeting Fph enzymes.
• To identify potential diagnostic and therapeutic agents against S. aureus.

Main Methods:

• Utilized mRNA display with a genetically encoded oxadiazolone (Ox) electrophile to screen diverse cyclic peptide libraries.
• Employed counter selections with wild-type (WT) and catalytic-dead FphB to enhance probe selectivity.
• Developed fluorescent probes based on selected cyclic peptides for FphB active site labeling.

Main Results:

• Identified potent and selective cyclic peptides that covalently inhibit Fph enzymes.
• Developed fluorescent probes that label the active site of FphB at single-digit nanomolar concentrations.
• Demonstrated successful labeling of FphB in live S. aureus bacteria.

Conclusions:

• Genetically encoded electrophiles in mRNA display are effective for generating covalent binding ligands.
• The developed fluorescent probes are potent and selective for FphB.
• These probes hold promise for the development of new diagnostics and therapeutics for S. aureus infections.