Lymphostatin: Structure of a large multi-functional virulence factor

Abstract

Enteropathogenic and Enterohaemorrhagic Escherichia coli are enteric pathogens of global importance and human infections can be life-threatening. Lymphostatin is a key virulence factor of these bacteria, being required for intestinal colonisation and a potent inhibitor of the mitogen- and antigen-activated proliferation of lymphocytes and proinflammatory responses. In some strains, it also mediates adherence to host cells and influences actin nucleation at sites of attachment. This 365 kDa protein requires glycosyltransferase and cysteine protease motifs for activity against lymphocytes, but high-resolution structural information has proven elusive and the molecular mechanisms by which it acts remain unclear. Here, we describe the structure of lymphostatin from the prototype O127:H6 enteropathogenic E. coli strain determined by electron cryo-microscopy. Our results reveal two glycosyltransferase domains, one PaTox-like protease domain, an ADP-ribosyltransferase domain, and a delivery domain. Long linkers act to hold these domains together. These linkers occlude the catalytic sites of the N-terminal glycosyltransferase and protease domains. In this dormant state, lymphostatin binds to HEK-293T cells, where it forms large clusters before being taken up and sequestered into cytosolic foci. With more functional domains than any other known large bacterial toxin, lymphostatin can be regarded as the multifunctional Swiss army knife of pathogenic Escherichia coli, enabling complex interactions with the host cells in different environments.