Fig. 2: The structure of Wzc^K540M determined by single-particle cryo-EM.

a The monomer of Wzc^K540M contains a cytoplasmic region (red), tyrosine-rich tail (yellow), two transmembrane helices (green) and the periplasmic region (blue, salmon and ochre). b A schematic representation of the structure of Wzc^K540M coloured as a The first residue of each structural block is shown. c Wzc^K540M forms an octamer. The transmembrane helices are not close-packed and create portals to a large central cavity, clearly visible in the space fill representation. The structure has both a periplasmic and a cytoplasmic ring. d The Wzc^K540M octamer viewed from the periplasm, reveals the periplasmic region also forms a ring-like arrangement. The central cavity is open to both the cytoplasm and lipid bilayer. Residues 65 to 84 could not be located experimentally and may occlude the entrance to the cavity from the periplasm. e The high quality of the EM map is illustrated by the holes for the aromatic residues. f The C-terminal tail from one monomer (coloured yellow and red) has residues labelled in normal text. Y717 is at the active site of the kinase domain from the other monomer (coloured cyan). Key kinase residues in the other monomer are labelled in bold italics. ADP and Mg²⁺ are shown and labelled. g With Y717 at the active site, Y715 sits in a pocket where it makes a hydrogen bond with E675 from the neighbouring subunit. A phosphorylated tyrosine at position i would be disfavoured by size and charge. Disruption of binding at position i could perturb binding at position i + 2, the active site. Thus phosphorylation of Y715 seems most likely to follow (not precede) phosphorylation of Y717. Residues from the kinase domain are labelled in italics.