Mechanism of catalysis, E2 recognition, and autoinhibition for the IpaH family of bacterial ubiquitin ligases.
A new study from the Sicheri lab at the Lunenfeld–Tanenbaum Research Institute (LTRI) provides insight, exploring how bacteria subvert host immunity.

In their article in Proceedings of the National Academy of Science (USA) journal titled “Mechanism of catalysis, E2 recognition, and autoinhibition for the IpaH family of bacterial ubiquitin ligases,” a team of researchers from the LTRI have delved into the role of a protein modification termed ubiquitination, a process that is key to immune defense against infection. In three enzymatic steps, this pathway causes the attachment of a small protein, ubiquitin (Ub), to another protein and flags this newly ubiquitinated protein for a different fate which includes its rapid destruction. In other words, this process allows a host (such as a human) to recognize and destroy a pathogen (such as Shigella).

To avoid this disposal path, pathogens like Shigella secrete other proteins that interrupt or modify the host’s ubiquitination machinery. The IpaH family of proteins, present in Shigella genome, subvert host Ub by mimicking the functions of an E3 ligase, one of the enzymes that catalyzes the attachment of ubiquitin to the target protein.

In their study, authors Alexander Keszei, a PhD candidate in the department of Molecular Genetics at the University of Toronto, and Dr. Frank Sicheri, senior investigator at the LTRI, explored an additional way in which IpaH interrupts Ub signaling. They found compelling evidence that IpaH proteins restrict the amounts of available “charged” E2 enzymes. Charged E2s are produced in the second step of ubiquitination and are requisite for E3 enzymes to catalyze the final step of ubiquitination. Hence, suppressing the levels of charged E2 proteins is a new way in which a pathogen can interrupt host ubiquitination.

Expanding upon this novel discovery, Keszei and Sicheri explored the mechanism involved in IpaH’s control of E2 levels and thus ubiquitination by identifying the sites on IpaH which bind target proteins in order to modify them. They demonstrated how one site in the protein’s NEL (Novel E3 Ligase) domain acts in a process termed aminolysis, while another is crucial in binding E2. This research also examines the function of another of the protein’s domains, the LRR (leucine–rich repeat) domain. It determines that this domain controls E3 ligase function, both by regulating its own E3 function as encoded by the protein’s NEL domain, as well as by controlling the E3 function of its host.

This study is an example of how modern structural biology, coupled with biochemistry and molecular biology can tease apart the means by which pathogenic bacteria can cause disease. In dissecting how IpaH works, the research represents a step toward to understanding how pathogens overcome their host’s immune responses. Discoveries such as these can also provide alternative strategies to combat the rising crisis ofantibiotic resistance. Current antibiotics work by targeting and killing infectious bacteria, but new drugs are continually needed as the microbes evolve mechanisms to neutralize the effects of the drugs. Research that provides an understanding of how bacteria subvert host immunity could provide a new kind of therapy that specifically targets bacteria’s infectious capability. As Alexander Keszei noted, “this could be a way of disarming Shigella.”