The orientation and stability of the GPCR-Arrestin complex in a lipid bilayer

Abstract

G protein-coupled receptors (GPCRs) constitute a large family of membrane proteins that plays a key role in transmembrane signal transduction and draw wide attention since it was discovered. Arrestin is a small family of proteins which can bind to GPCRs, block G protein interactions and redirect signaling to G-protein-independent pathways. The detailed mechanism of how arrestin interacts with GPCR remains elusive. Here, we conducted molecular dynamics simulations with coarse-grained (CG) and all-atom (AA) models to study the complex structure formed by arrestin and rhodopsin, a prototypical GPCR, in a POPC bilayer. Our results indicate that the formation of the complex has a significant impact on arrestin which is tightly anchored onto the bilayer surface, while has a minor effect on the orientation of rhodopsin in the lipid bilayer. The formation of the complex induces an internal change of conformation and flexibility in both rhodopsin and arrestin, mainly at the binding interface. Further investigation on the interaction interface identified the hydrogen bond network, especially the long-lived hydrogen bonds, and the key residues at the contact interface, which are responsible for stabilizing the complex. These results help us to better understand how rhodopsin interacts with arrestin on membranes, and thereby shed lights on arrestin-mediated signal transduction through GPCRs.

Figure 1

The initial system for molecular dynamics simulations. The gray surface represents the POPC lipid bilayer. The green cartoon indicates rhodopsin and the cyan cartoon indicates arrestin. The blue and red spheres represent Na⁺ and CL⁻ respectively. Water molecules were filled in the whole simulation box but are not shown here for clarity.

Figure 2

The first principal axis of the rhodopsin-arrestin complex, rhodopsin alone and arrestin alone. Red arrow represents the first principal axis. Brown spheres indicates the position of PO₄ in the POPC bilayers (a) The first principal axis of the rhodopsin-arrestin complex in a POPC bilayer. (b) The first principal axis of the standalone rhodopsin, where rhodopsin was embedded into a POPC bilayer. (c) The first principal axis of the standalone arrestin near a POPC bilayer.

Figure 3

The orientation of the complex orientation in a POPC bilayer. The orientation of the complex is represented by the angle between the first principal axis of the rhodopsin-arrestin complex structure and the Z axis of the simulation box. (a) All the data from ten CG complex trajectories were plotted with black lines. Data from the ten CG complex without ELN were plotted with red lines. Blue line indicates the results from the AA MD simulation. (b–d) The angle probability distribution of the CG complex, AA complex and CG complex without ELN in MD, respectively.

Figure 4

The orientation of rhodopsin and arrestin before and after the complex formation. All data from rhodopsin alone or arrestin alone were plotted with black lines, data from rhodopsin in complex or arrestin in complex were plotted with red lines. (a) The rhodopsin orientation change was subtle upon the formation of the complex. (b) The probability distribution of the angle between the first principal axis of rhodopsin and the Z-axis, before (top) and after (bottom) the complex formation. (c) The arrestin orientation was limited to a much smaller range upon the complex formation. (d) same as (b), but for arrestin.

Figure 5

Principal component analysis (PCA) and RMSF change of rhodopsin and arrestin. (a) The results of PCA. (b) Two extreme conformations of rhodopsin and arrestin as determined by PCA. The transparent structure represents the conformation in complex, and the opaque structure represents the standalone conformation. (c) The RMSF change of the residues of rhodopsin and arrestin upon the complex formation. (d) The colored complex structure according to the RMSF change of the residues with the blue-white-red code.

Figure 6

Details of the two types hydrogen bonds. Red dashed lines represent hydrogen bonds. Water was shown with the CPK model. (a) T242_(rhodopsin) side chain and D83_(arrestin) side chain; T242_(rhodopsin) side chain and D83_(arrestin) main chain. (b) Q237_(rhodopsin) side chain and R82_(arrestin) side chain. (c) V138_(rhodopsin) main chain and Y68_(arrestin) side chain; R135_(rhodopsin) side chain and M76_(arrestin) main chain. (d) F146_(rhodopsin) main chain and K142_(arrestin) side chain; F148_(rhodopsin) main chain and K142_(arrestin) side chain. (e) the hydrogen bonds between E249_(rhodopsin) and L78_(arrestin); the hydrogen bonds between V138_(rhodopsin) and T79_(arrestin); the hydrogen bonds between N145_(rhodopsin) and Y68_(arrestin). (f) the hydrogen bonds between F146_(rhodopsin) and D139_(arrestin); the hydrogen bonds between R147_(rhodopsin) and Q138_(arrestin). (g) the hydrogen bonds between Q236_(rhodopsin) and L250_(arrestin). (h) the hydrogen bonds between K66_(rhodopsin) and D72_(arrestin).

Figure 7

The residue contact map at the rhodopsin-arrestin interface. (a) Site hb1 represents type A hydrogen bonds in Fig. 6c; hb2 represents type A hydrogen bonds in Fig. 6a; hb3 represents type A hydrogen bonds in Fig. 6d; and hb4 represents type B hydrogen bonds in Fig. 6f. (b–d) Electrostatic interaction sites at the complex interface. (e–g) Hydrophobic contacts at the complex interface.