Molecular Mechanism of Membrane Binding of the GRP1 PH Domain

Abstract

The pleckstrin homology (PH) domain of the general receptor of phosphoinositides 1 (GRP1) protein selectively binds to a rare signaling phospholipid, phosphatidylinositol (3,4,5)-trisphosphate (PIP3), in the membrane. The specific PIP3 lipid docking of GRP1 PH domain is essential to protein cellular function and is believed to occur in a stepwise process, electrostatic-driven membrane association followed by the specific PIP3 binding. By a combination of all-atom molecular dynamics (MD) simulations, coarse-grained analysis, electron paramagnetic resonance (EPR) membrane docking geometry, and fluorescence resonance energy transfer (FRET) kinetic studies, we have investigated the search and bind process in the GRP1 PH domain at the molecular scale. We simulated the two membrane binding states of the GRP1 PH domain in the PIP3 search process, before and after the GRP1 PH domain docks with the PIP3 lipid. Our results suggest that the background anionic phosphatidylserine lipids, which constitute around one-fifth of the membrane by composition, play a critical role in the initial stages of recruiting protein to the membrane surface through non-specific electrostatic interactions. Our data also reveal a previously unseen transient membrane association mechanism that is proposed to enable a two-dimensional “hopping” search of the membrane surface for the rare PIP3 target lipid. We further modeled the PIP3-bound membrane–protein system using the EPR membrane docking structure for the MD simulations, quantitatively validating the EPR membrane docking structure and augmenting our understanding of the binding interface with atomic-level detail. Several observations and hypotheses reached from our MD simulations are also supported by experimental kinetic studies.

Phosphatidylserine in the brain: Metabolism and function

“…The C 2 A domain binds to PS and the C 2 B domain binds to inositol phospholipids [166]. GRP1, a general receptor for phosphoinositides, transiently associates with the membrane through the binding between the PH domain and PS, which enables a two-dimensional search for its PIP 3 target [167]. …”

Allosteric regulation of Arf GTPases and their GEFs at the membrane interface

“…53 Biophysical and molecular dynamics simulations suggest that the PH domain and the polybasic region bind additional acidic lipids at non-canonical binding sites. 54,55 Whereas the activities and specificities of these 3 subfamilies differ broadly in solution, all are highly potent at activating both Arf1 and Arf6 in the presence of membranes, including EFA6 which is not active on Arf1 in solution. 29,46,49,[56][57][58] These stark differences highlight the importance to assess specificity using membranes and myristoylated Arf GTPases, as this is often the starting point for building models of the biology of the proteins being studied.…”

Interplay between phosphoinositide lipids and calcium signals at the leading edge of chemotaxing ameboid cells

“…If physiological conditions produce membrane regions containing both PI(4,5)P 2 clusters and PS, such regions could exhibit increased affinity for cPKC C2 domains that require both PI(4,5)P 2 and PS for optimal target membrane affinity and specificity (Evans and Falke, 2007; Corbalan-Garcia et al, 2007; Corbin et al, 2007; Evans et al, 2006). Similarly, PH domains require both PI(3,4,5)P 3 and PS for high affinity, rapid target membrane acquisition (Corbin et al, 2004; Knight and Falke, 2009; Lai et al, 2013), thus their targeting reactions could be facilitated by regions containing both PI(3,4,5)P 3 clusters and PS.…”