Molecular basis for polysaccharide recognition and modulated ATP hydrolysis by the O antigen ABC transporter

• Published in: Nature communications Vol. 13; no. 1; pp. 5226 - 14
• Main Authors: Spellmon, Nicholas, Muszyński, Artur, Górniak, Ireneusz, Vlach, Jiri, Hahn, David, Azadi, Parastoo, Zimmer, Jochen
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 05.09.2022; Nature Publishing Group; Nature Portfolio
• Subjects: 101/28; 631/326/421; 631/45/221; 631/45/612/1237; 631/535/1258/1259; ABC transporter; ABC transporters; Adenosine triphosphate; Antigens; BASIC BIOLOGICAL SCIENCES; Binding; Biosynthesis; Carbohydrates; Chemical modification; Cryoelectron microscopy; Crystal structure; Cytosol; Domains; Electron microscopy; Glycobiology; Humanities and Social Sciences; Lipids; Lipopolysaccharides; Mannose; Membrane proteins; multidisciplinary; Nucleotides; O antigen; Pathogens; Periplasm; Polymers; Polysaccharides; Recognition; Rhamnose; Saccharides; Science; Science (multidisciplinary); Site-directed mutagenesis; Translocation
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-022-32597-2

O antigens are ubiquitous protective extensions of lipopolysaccharides in the extracellular leaflet of the Gram-negative outer membrane. Following biosynthesis in the cytosol, the lipid-linked polysaccharide is transported to the periplasm by the WzmWzt ABC transporter. Often, O antigen secretion requires the chemical modification of its elongating terminus, which the transporter recognizes via a carbohydrate-binding domain (CBD). Here, using components from A. aeolicus , we identify the O antigen structure with methylated mannose or rhamnose as its cap. Crystal and cryo electron microscopy structures reveal how WzmWzt recognizes this cap between its carbohydrate and nucleotide-binding domains in a nucleotide-free state. ATP binding induces drastic conformational changes of its CBD, terminating interactions with the O antigen. ATPase assays and site directed mutagenesis reveal reduced hydrolytic activity upon O antigen binding, likely to facilitate polymer loading into the ABC transporter. Our results elucidate critical steps in the recognition and translocation of polysaccharides by ABC transporters. Gram negative microbes strategically equip lipopolysaccharides with O antigens to expand the extracellular envelope. Here, the authors examine how the O antigen is captured by a special class of ABC transporters containing a carbohydrate-binding module accessory.