Membrane-Permeable Antibacterial Enzyme against Multidrug-Resistant Acinetobacter baumannii

Bacteriophage endolysins (lysins, or murein hydrolases) are enzymes that bacteriophages utilize to degrade the cell wall peptidoglycans (PG) and subsequently disintegrate bacterial cells from within. Due to their muralytic activity, lysins are considered as potential candidates to battle against antibiotic resistance. However, most lysins in their native form lack the capability of trespassing the outer membrane (OM) of Gram-negative (G-ve) bacteria. To turn the bacteriophage enzymes into antibacterial weapons against G-ve bacteria, endowing these enzymes the capability of accessing the PG substrate underneath the OM is critical. Here we show that fusing a membrane-permeabilizing peptide CeA at the C-terminus of a muralytic enzyme LysAB2 renders a two-step mechanism of bacterial killing and increases the activity of LysAB2 against the multidrug resistant Acinetobacter baumannii by up to 100 000-folds. The engineered LysAB2, termed LysAB2-KWK here, also shows remarkable activity against A. baumannii at the stationary phase and a prominent capability to disrupt biofilm formation. In addition, the enzyme shows a broad antibacterial spectrum against G-ve bacteria, a decent tolerance to serum, and a prolonged storage life. LysAB2-KWK rescues the larva of the greater wax moth Galleria mellonella from A. baumannii infection through systemic administration. Altogether, our work equips a globular lysin with OM permeabilization activity to enable effective killing of G-ve bacteria, reveals the critical role of the C-terminus of a globular lysin in the antibacterial activity, and points toward a viable route to engineer globular lysins as antibacterial enzymes for potential clinical use against multidrug resistant G-ve bacteria.