The <scp>I</scp>ntimin periplasmic domain mediates dimerisation and binding to peptidoglycan

<jats:title>Summary</jats:title><jats:p><jats:styled-content style="fixed-case">I</jats:styled-content>ntimin and <jats:styled-content style="fixed-case">I</jats:styled-content>nvasin are prototypical inverse (<jats:styled-content style="fixed-case">T</jats:styled-content>ype <jats:styled-content style="fixed-case">V</jats:styled-content>e) autotransporters and important virulence factors of enteropathogenic <jats:styled-content style="fixed-case"><jats:italic>E</jats:italic></jats:styled-content><jats:italic>scherichia coli</jats:italic> and <jats:styled-content style="fixed-case"><jats:italic>Y</jats:italic></jats:styled-content><jats:italic>ersinia spp</jats:italic>. respectively. In addition to a <jats:styled-content style="fixed-case">C</jats:styled-content>‐terminal extracellular domain and a β‐barrel transmembrane domain, both proteins also contain a short <jats:styled-content style="fixed-case">N</jats:styled-content>‐terminal periplasmic domain that, in <jats:styled-content style="fixed-case">I</jats:styled-content>ntimin, includes a lysin motif (<jats:styled-content style="fixed-case">LysM</jats:styled-content>), which is thought to mediate binding to peptidoglycan. We show that the periplasmic domain of <jats:styled-content style="fixed-case">I</jats:styled-content>ntimin does bind to peptidoglycan both <jats:italic>in vitro</jats:italic> and <jats:italic>in vivo</jats:italic>, but only under acidic conditions. We were able to determine a dissociation constant of 0.8 μM for this interaction, whereas the <jats:styled-content style="fixed-case">I</jats:styled-content>nvasin periplasmic domain, which lacks a <jats:styled-content style="fixed-case">LysM</jats:styled-content>, bound only weakly <jats:italic>in vitro</jats:italic> and failed to bind peptidoglycan <jats:italic>in vivo</jats:italic>. We present the solution structure of the <jats:styled-content style="fixed-case">I</jats:styled-content>ntimin <jats:styled-content style="fixed-case">LysM</jats:styled-content>, which has an additional α‐helix conserved within inverse autotransporter <jats:styled-content style="fixed-case">L</jats:styled-content>ys<jats:styled-content style="fixed-case">M</jats:styled-content>s but lacking in others. In contrast to previous reports, we demonstrate that the periplasmic domain of <jats:styled-content style="fixed-case">I</jats:styled-content>ntimin mediates dimerisation. We further show that dimerisation and peptidoglycan binding are general features of <jats:styled-content style="fixed-case">LysM</jats:styled-content>‐containing inverse autotransporters. Peptidoglycan binding by the periplasmic domain in the infection process may aid in resisting mechanical and chemical stress during transit through the gastrointestinal tract.</jats:p>