mGBP7 interacting proteins in Toxoplasma gondii infection and biochemical characteristics of mGBP7

• Published in: The Journal of immunology (1950) 2019-05, Vol.202 (1_Supplement), p.127-127.8
• Main Authors: Legewie, Larissa, Loschwitz, Jennifer, Smits, Sander, Prescher, Martin, Wang, Xue, Steffens, Nora, Degrandi, Daniel, Strodel, Birgit, Schmitt, Lutz, Pfeffer, Klaus
• Format: Article
• Language: English
• ISSN: 0022-1767; 1550-6606
• DOI: 10.4049/jimmunol.202.Supp.127.8

Toxoplasma gondii (T. gondii) is an obligate intracellular parasite and the causative agent of toxoplasmosis. During host cell invasion, T. gondii forms an unique membranous compartment called the parasitophorous vacuole (PV). Members of the murine guanylate binding protein (mGBP) family localize around the PV leading to its disruption, but the underlying molecular mechanisms are poorly understood. The aim of this project is to provide a better understanding of the biochemistry of mGBPs as well as to elucidate the molecular mechanisms by which this IFNγ-inducible protein family is able to attack the parasite directly. The GTPase activity of mGBP7 and its oligomerization status were analyzed using a colorimetric assay and multi-angle light scattering (MALS). The structure model of mGBP7 was obtained using I-TASSER homology modeling. For the identification of potential mGBP7 interaction partners co-immunoprecipitation (IP) and mass spectrometry (MS) experiments were performed. The obtained biochemical data for mGBP7 display a GTPase activity in the μM range and a positive cooperativity of GTP hydrolysis. Furthermore, the MALS results indicate a transient oligomer formation independent of the presence of nucleotides. Homology modeling reveals an extended C-terminal domain for mGBP7 probably responsible for its oligomerization pattern. The initial mGBP7 IP-MS results offer hints about potential mGBP7 interaction partners and indicate that mGBP7 is ubiquitinated and ISGylated even in the absence of T. gondii infection. The biochemical characterization of mGBP7 as well as the identification of potential mGBP7 interacting proteins will provide new insights into the dynamic interactions at the interface of parasite and host.