A biosynthetic strategy for re-engineering the Staphylococcus aureus cell wall with non-native small molecules

Area of Science:

• Microbiology
• Synthetic Biology
• Biochemistry

Background:

• Staphylococcus aureus is a significant Gram-positive bacterial pathogen posing public health risks.
• The bacterial cell wall, particularly peptidoglycan, is crucial for bacterial structure and survival.
• Modifying the bacterial cell wall offers potential for new therapeutic and diagnostic strategies.

Purpose of the Study:

• To develop a method for covalently re-engineering the cell wall of Staphylococcus aureus.
• To incorporate non-native small molecules into the S. aureus cell wall using endogenous enzymes.
• To demonstrate the utility of engineered cell wall components for further functionalization.

Main Methods:

• Utilized the endogenous bacterial enzyme sortase A (SrtA) for covalent modification.
• Incubated wild-type S. aureus with rationally designed SrtA substrates containing functional molecular handles (fluorescein, biotin, azide).
• Employed epifluorescence and electron microscopy, biochemical extraction, and mass spectrometry for characterization.

Main Results:

• Successfully demonstrated covalent incorporation of fluorescein, biotin, and azide into the S. aureus cell wall peptidoglycan.
• Confirmed the presence and location of incorporated molecules using various analytical techniques.
• Showcased the utility of the incorporated azide handle for surface functionalization via click chemistry (azide-alkyne cycloaddition).

Conclusions:

• This study presents the first example of cell wall engineering in Staphylococcus aureus or any pathogenic Gram-positive bacteria.
• The developed method enables the covalent attachment of functional molecular handles to the bacterial cell wall.
• This approach holds significant potential for diverse applications in microbiology and biotechnology.