Architecture of a channel-forming O-antigen polysaccharide ABC transporter

O-antigens are cell surface polysaccharides of many Gram-negative pathogens that aid in escaping innate immune responses.(1) A widespread O-antigen biosynthesis mechanism involves the synthesis of the lipid-anchored polymer on the inner membrane’s (IM’s) cytosolic face, followed by transport to the periplasmic side where it is ligated to the lipid A core to complete a lipopolysaccharide (LPS) molecule(2). The O antigen’s transport to the periplasm is mediated by an ATP-binding cassette (ABC) transporter, called Wzm/Wzt, Extended Data Fig. 1. We present the crystal structure of the Wzm/Wzt homolog from Aquifex aeolicus in an open conformation. The transporter forms a transmembrane (TM) channel sufficiently wide to accommodate a linear polysaccharide. It’s nucleotide binding domain (NBD) and a periplasmic loop form ‘gate helices’ at the cytosolic and periplasmic membrane interfaces, likely serving as substrate entry and exit points. Site-directed mutagenesis of the gates impairs in vivo O antigen secretion in the E. coli prototype. Combined with a closed structure of the isolated NBDs, our structural and functional analyses suggest a processive O antigen translocation mechanism, which stands in contrast to the classical alternating access mechanism of ABC transporters.