Domain motions, dimerization, and membrane interactions of the murine guanylate binding protein 2

Abstract

Guanylate-binding proteins (GBPs) are a group of interferon-inducible GTPases that are essential components of cell-autonomous immunity against intracellular pathogens. They restrict the replication of intracellular pathogens by targeting and lysing the membrane of the pathogen-containing vacuole. The aim of our work is to provide a molecular-level understanding of how the GBPs destroy the parasitophorous vacuole membrane. To this end, we use molecular dynamics (MD) simulations to elucidate the domain motions of GBPs and the effects on these motions following GTP binding, GBP multimerization, and membrane binding, assuming that each of these steps plays important roles in the defense mechanism. Here, we present our results for the murine GBP2, one of the eleven GBPs present in mice, obtained from standard and replica exchange MD simulations with an accumulated simulation time of 30 µs. The main findings are that mGBP2 features a large-scale hinge motion in its M/E domain which is not hugely affected by either GTP binding, geranylgeranylation, homodimer formation, or lipid-membrane binding. Our prognosis is that this hinge motion has the potential to destroy the membrane following the formation of supramolecular mGBP2 complexes on the membrane surface.

Integrative modeling of guanylate binding protein dimers

“…Before establishing dimer models of the three GBPs under investigation, we compare the dynamics of their monomers, on the one hand to further elucidate possible differences between hGBP1, mGBP2, and mGBP7, and on the other hand to have the monomer dynamics as a reference for the dynamics of the proteins when being oligomerized. The dynamics of hGBP1 and mGBP2 was discussed in detail in our previous works, 22,23 while the dynamics of mGBP7 is being unraveled in this study. In all three cases, Hamiltonian replica exchange molecular dynamics (HREMD) simulations were applied to the monomeric proteins.…”

“…Using bioinformatic/phylogenetic methods, seven human GBPs (hGBP1 to hGBP7) and 11 murine GBPs (mGBP1 to mGBP11) have been identified. 15,21 In this work, we will concentrate on hGBP1, mGBP2, and mGBP7, continuing our previous studies of hGBP1, 22 mGBP2, 23 and mGBP7. 24 The GBPs exhibit three domains (Fig.…”

“…For mGBP2 we had shown that the smaller HREMD setup was sufficient as no further protein states were sampled in a 40 × 400 ns HREMD simulation. 23 For the analysis, we considered the protein conformations collected in the HREMD target replica that had no modifications applied to the potential energy function of either hGBP1, mGBP2, or mGBP7.…”

“…were prepared for the simulations as previously described, which includes the parametrization of GTP and the geranylgeranyl group. [22][23][24] The hGBP1 dimer was created using MMalign (https://zhanglab.ccmb.med.umich.edu/MM-align/ 58 ). For all MD simulations, the Amber99SB*-ILDNP force field [78][79][80] was used for modeling the proteins, the TIP3P model was used for water, 81 and GROMACS 2016 82,83 was employed for running the simulations.…”

“…To identify the role of GTP and the CT tail, two further systems, mGBP7 GTP and mGBP7 holo were created, using an identical approach as described on our work of mGBP2. 23 The protein was always placed in a rectangle simulation box of 10 nm × 10 nm × 18.0 nm dimensions, ~56,000 water molecules were added, as well as 10 (apo), 13 (GTP), or 13 Na + (holo) for the neutralization of the systems, resulting in a total number of ~178,000 atoms. A similar HREMD protocol as applied in our study of mGBP2 dynamics was used and is described in detail in the Supplementary Information.…”