Solution structure and dynamics of the mitochondrial-targeted GTPase-activating protein (GAP) VopE by an integrated NMR/SAXS approach

Author:

Smith, Kyle P [ORCID]

Lee, Woonghee [ORCID]

Tonelli, Marco

Lee, Yeongjoon

Light, Samuel H

Cornilescu,Gabriel

Chakravarthy, Srinivas
Author Address

Current, Xilio Therapeutics, Waltham, MA, USA., Former, Department of Cell & Developmental Biology, Northwestern University Chicago, IL, USA., Department of Chemistry, University of Colorado-Denver, Denver, CO, USA., National Magnetic Resonance Facility at Madison, Department of Biochemistry, University of Wisconsin-Madison, Madison, WI, USA., Department of Microbiology, University of Chicago, Chicago, IL, USA., Advanced Technology Research Facility, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., National Cancer Institute, National Institutes of Health, Frederick, MD, USA., Biophysics Collaborative Access Team, Argonne National Laboratory, Argonne, IL, USA.,

Abstract:

The bacterial pathogen Vibrio cholerae use a type III secretion system to inject effector proteins into a host cell. Recently, a putative Toxic GTPase Activating Protein (ToxGAP) called VopE was identified as a T3SS substrate and virulence factor that affected host mitochondrial dynamics and immune response. However, biophysical and structural characterization has been absent. Here, we describe solution NMR structure of the putative GAP domain (73-204) of VopE. Using SEC-SAXS and RDC data, we restrained the MD process to efficiently determine the overall fold and improve the quality of the output calculated structures. Comparing the structure of VopE with other ToxGAP's revealed a similar overall fold with several features unique to VopE. Specifically, the "Bulge 1", a1 helix, and noteworthy "backside linker" elements on the N-terminus are dissimilar to the other ToxGAP's. By using NMR relaxation dispersion experiments, we demonstrate that these regions undergo motions on a >6 s-1 timescale. Based on the disposition of these mobile regions relative to the putative catalytic arginine residue, we hypothesize the protein may undergo structural changes to bind cognate GTPases. This article is protected by copyright. All rights reserved. © 2022 The Protein Society.

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