Membrane-binding mechanism of the EEA1 FYVE domain revealed by multi-scale molecular dynamics simulations

Abstract

Early Endosomal Antigen 1 (EEA1) is a key protein in endosomal trafficking and is implicated in both autoimmune and neurological diseases. The C-terminal FYVE domain of EEA1 binds endosomal membranes, which contain phosphatidylinositol-3-phosphate (PI(3)P). Although it is known that FYVE binds PI(3)P specifically, it has not previously been described of how FYVE attaches and binds to endosomal membranes. In this study, we employed both coarse-grained (CG) and atomistic (AT) molecular dynamics (MD) simulations to determine how FYVE binds to PI(3)P-containing membranes. CG-MD showed that the dominant membrane binding mode resembles the crystal structure of EEA1 FYVE domain in complex with inositol-1,3-diphospate (PDB ID 1JOC). FYVE, which is a homodimer, binds the membrane via a hinge mechanism, where the C-terminus of one monomer first attaches to the membrane, followed by the C-terminus of the other monomer. The estimated total binding energy is ~70 kJ/mol, of which 50–60 kJ/mol stems from specific PI(3)P-interactions. By AT-MD, we could partition the binding mode into two types: (i) adhesion by electrostatic FYVE-PI(3)P interaction, and (ii) insertion of amphipathic loops. The AT simulations also demonstrated flexibility within the FYVE homodimer between the C-terminal heads and coiled-coil stem. This leads to a dynamic model whereby the 200 nm long coiled coil attached to the FYVE domain dimer can amplify local hinge-bending motions such that the Rab5-binding domain at the other end of the coiled coil can explore an area of 0.1 μm2 in the search for a second endosome with which to interact.

Specific interactions of peripheral membrane proteins with lipids: what can molecular simulations show us?

“…CG MD involve approximating the chemical landscape, as described previously. In cases with available experimental data, however, good consistency has reported between experimental energies and energies obtained with PMF using Martini 2 (16,123). AA MD is a more accurate description of the system, but it is more challenging to obtain convergence (38).…”

“…Electrostatic interactions can be mediated via positive ions (typically calcium) that bridge negatively-charged patches on the PMP with negative lipids (8,9) (Figure 2B). Alternative forms of interaction exist, such as insertion of hydrophobic loops (10)(11)(12)(13)(14)(15)(16) or hydrophobic helices (17) into the membrane, cation-π interactions between aromatic residues and choline headgroups (18,19), or covalent attachment to a lipid whose hydrophobic tail is inserted into the lipid bilayer (20) (Figure 2). (21).…”

“…That is, avidity must be considered as affinities between single molecules are not trivially additive. Binding of multiple PIP 2 were likewise observed for different membrane-binding C2 domains [ 21 ], for Ebola virus matrix protein VP40 [ 121 ] and for Brag2 [ 122 ], and multiple PI(4)P were found to bind the FYVE domain of early endosome antigen 1 [ 16 ].…”

“…The umbrella sampling PMF method has been applied to a number of PMPs, including studies of PH domains [ 94 , 120 ], C2 domains [ 21 ], the FYVE domain of early endosome antigen 1 [ 16 ] and the related lipidated GTPase Rab5 [ 23 ], which are both involved in early endosome recruiting. An example for a C2 domain is given in Figure 4 B.…”

“…( C ) Loop insertion . The FYVE domain of EEA1 (zinc ions from the four zinc fingers in blue) has a partly hydrophobic loop (V44 shown in inset) which plays a part in the membrane binding [ 16 ]. ( D ) Helix insertion .…”