Integrative modeling of guanylate binding protein dimers

Abstract

Guanylate‐binding proteins (GBPs) are essential interferon‐γ‐activated large GTPases that play a crucial role in host defense against intracellular bacteria and parasites. While their protective functions rely on protein polymerization, our understanding of the structural intricacies of these multimerized states remains limited. To bridge this knowledge gap, we present dimer models for human GBP1 (hGBP1) and murine GBP2 and 7 (mGBP2 and mGBP7) using an integrative approach, incorporating the crystal structure of hGBP1's GTPase domain dimer, crosslinking mass spectrometry (XL‐MS), small‐angle X‐ray scattering (SAXS), protein‐protein docking, and molecular dynamics (MD) simulations. Our investigation begins by comparing the protein dynamics of hGBP1, mGBP2, and mGBP7. We observe that the M/E domain in all three proteins exhibits significant mobility and hinge motion, with mGBP7 displaying a slightly less pronounced motion but greater flexibility in its GTPase domain. These dynamic distinctions can be attributed to variations in the sequences of mGBP7 and hGBP1/mGBP2, resulting in different dimerization modes. Unlike hGBP1 and its close ortholog mGBP2, which exclusively dimerize through their GTPase domains, we find that mGBP7 exhibits three equally probable alternative dimer structures. The GTPase domain of mGBP7 is only partially involved in its dimerization, primarily due to an accumulation of negative charge, allowing mGBP7 to dimerize independently of GTP. Instead, mGBP7 exhibits a strong tendency to dimerize in an antiparallel arrangement across its stalks. The results of this work go beyond the sequence‐structure‐function relationship, as the sequence differences in mGBP7 and mGBP2/hGBP1 do not lead to different structures, but to different protein dynamics and dimerization. The distinct GBP dimer structures are expected to encode specific functions crucial for disrupting pathogen membranes.This article is protected by copyright. All rights reserved.