The Toxin Complex (Tc) superfamily consists of toxin translocases that contribute to the targeting, delivery, and cytotoxicity of certain pathogenic Gram-negative bacteria. Membrane receptor targeting is driven by the A-subunit (TcA), which comprises IgG-like receptor binding domains (RBDs) at the surface. To better understand XptA2, an insect specific TcA secreted by the symbiont X. nematophilus from the intestine of entomopathogenic nematodes, we determined structures by X-ray crystallography and cryo-EM. Contrary to a previous report, XptA2 is pentameric. RBD-B exhibits an indentation from crystal packing that indicates loose association with the shell and a hotspot for possible receptor binding or a trigger for conformational dynamics. A two-fragment XptA2 lacking an intact linker achieved the folded pre-pore state like wild type (wt), revealing no requirement of the linker for protein folding. The linker is disordered in all structures, and we propose it plays a role in dynamics downstream of the initial pre-pore state.
|Journal||International Journal of Molecular Sciences|
|State||Published - Sep 2023|
Bibliographical noteFunding Information:
This research was supported by a UAB competitive COVID-19 relief award and start-up funds to S.G.A.
CryoEM data was collected at the Baylor College of Medicine CryoEM ATC, which includes equipment purchased under the support of CPRIT Core Facility Award RP190602. EM data collection from the FEI Polara was conducted at the Center for Structural Biology Cryo-EM Facility at Vanderbilt University. The XptA2 2-fragment data was collected in the UAB cryo-EM core, and the DSF data was collected in the UAB X-ray Crystallography Core Facility supported by the Comprehensive Cancer Center Support Grant (NIH grant # 2P30CA013148-49). C.D.R. is supported by the National Institute of Health, United States, grant 1K99AI166116. We thank Joel Sheets for the initial purification of XptA2 for crystallization. This work is based on C.L.M.’s dissertation submitted to fulfill, in part, the requirements for the degree of Doctor of Philosophy at the University of Alabama at Birmingham.
© 2023 by the authors.
- X-ray crystallography
- toxin translocase
ASJC Scopus subject areas
- Molecular Biology
- Computer Science Applications
- Physical and Theoretical Chemistry
- Organic Chemistry
- Inorganic Chemistry