Abstract Target protection proteins bind to antibiotic targets and confer resistance to the host organism. One class of such proteins, termed a ntibiotic re sistance (ARE) ATP binding cassette (ABC) proteins of the F-subtype (ARE ABCFs), are widely distributed throughout Gram-positive bacteria and bind the ribosome to alleviate translational inhibition by antibiotics that target the large ribosomal subunit. Using single-particle cryo-EM, we have solved the structure of ARE ABCF–ribosome complexes from three Gram-positive pathogens: Enterococcus faecalis LsaA, Staphylococcus haemolyticus VgaA LC and Listeria monocytogenes VgaL. Supported by extensive mutagenesis analysis, these structures enable a comparative approach to understanding how these proteins mediate antibiotic resistance on the ribosome. We present evidence of mechanistically diverse allosteric relays converging on a few peptidyltransferase center (PTC) nucleotides, and propose a general model of antibiotic resistance mediated by these ARE ABCFs.

Structural basis of resistance to lincosamide, streptogramin A, and pleuromutilin antibiotics by ABCF ATPases in Gram-positive pathogens