Chemically Induced Cell Wall Stapling in Bacteria

Abstract

Transpeptidation reinforces the structure of cell-wall peptidoglycan, an extracellular heteropolymer that protects bacteria from osmotic lysis. The clinical success of transpeptidase-inhibiting β-lactam antibiotics illustrates the essentiality of these cross-linkages for cell-wall integrity, but the presence of multiple, seemingly redundant transpeptidases in many species makes it challenging to determine cross-link function. Here, we present a technique to link peptide strands by chemical rather than enzymatic reaction. We employ biocompatible click chemistry to induce triazole formation between azido- and alkynyl-d-alanine residues that are metabolically installed in the peptidoglycan of Gram-positive or Gram-negative bacteria. Synthetic triazole cross-links can be visualized using azidocoumarin-d-alanine, an amino acid derivative that undergoes fluorescent enhancement upon reaction with terminal alkynes. Cell-wall stapling protects Escherichia coli from treatment with the broad-spectrum β-lactams ampicillin and carbenicillin. Chemical control of cell-wall structure in live bacteria can provide functional insights that are orthogonal to those obtained by genetics.

Keywords: antibiotic; bacteria; cell wall; click chemistry; cross-linking; d-amino acid; metabolic labeling; peptidoglycan; transpeptidase.