How bacteria fight back against promising antimicrobial peptide

Antimicrobial peptides have potential in antibiotic drug development, including possible uses in combination with other antibiotics for infections that are difficult to treat. Scientists have shown that the peptide TAT-RasGAP317-326, originally developed as an anticancer compound, inhibits E. coli and Staphylococcus aureus, among other bacteria. The peptide contains residues 317-326 of the Ras GTPase-activating protein, or RasGAP, with an attached N-terminal cell-penetrating sequence from the HIV transactivator of transcription, or TAT, protein, and will be called TAT-RasGAP in this article for simplicity. Maria Georgieva at the University of Lausanne Hospital Center and a team in Switzerland performed a resistance selection experiment over 20 passages to obtain an E. coli strain resistant to TAT-RasGAP to identify mutations that could elucidate this peptide’s mechanism of action. In a recent Journal of Biological Chemistry article, they showed that a mutation in BamA, an outer membrane protein critical for the insertion of other membrane proteins, helped block the peptide’s antimicrobial activity.

David S. Goodsell, RCSB Protein Data Bank, via Wikimedia Commons

Illustration of a cross section of an E. coli cell. The cell wall is shown in green, the genome in yellow, DNA-binding proteins in tan and orange and ribosomes in purple.

The authors traced the mutation that protects E. coli from TAT-RasGAP to a negatively charged loop in BamA that extends into the extracellular space. The mutation changes a residue from a negative to a neutral charge. The authors hypothesized that the positively charged TAT-RasGAP may interact with this negatively charged loop for cell entry, and a negative-to-neutral mutation could have developed in the resistant strain to block this electrostatic interaction. Modeling and molecular dynamics indicated that BamA’s negatively charged loop likely interacts with the peptide.

However, further experiments showed that TAT-RasGAP does not produce the same  changes as known BamA inhibitors based on bacterial morphology viewed by brightfield microscopy and outer membrane protein quantification, indicating that BamA is unlikely inhibited by TAT-RasGAP. Future experiments will help resolve the full mechanism of action for TAT-RasGAP and could lead to novel antibiotics.