Conformational Ensembles from Experimental Data and Computer Simulations

Conformational Ensembles from Experimental Data

and Computer Simulations

Sunday Speaker Abstracts

Functional Dynamics of the Distal C-tail of Arrestin

Martha Sommer

, Ciara C.M. Lally

, Brian Bauer

Jana Selent

Pompeu Fabra University, Hospital del Mar Medical Research Institute, Barcelona,

Spain.

Institute of Medical Physics and Biophysics (CC2), Charité Medical University, Berlin,

Germany,

Arrestin proteins regulate the large and diverse family of G protein-coupled receptors (GPCRs).

Arrestins have an elongated structure consisting of two clam shell-like domains and a long C-

terminal tail (C-tail). In crystal structures of arrestin, the proximal C-tail is observed to interact

extensively with the N-domain, thereby stabilizing the basal state. However, the highly flexible

and negatively charged distal C-tail is not visible in the crystal structures. Displacement of the

entire C-tail by the phosphorylated receptor C-terminus is believed to activate arrestin for

receptor binding.

In this study, we have applied a combination of computational and biophysical methods in order

to investigate the structural dynamics of the arrestin distal C-tail. Molecular dynamics

simulations show the distal C-tail sampling a wide conformational space within the concave

surface of the N-domain, and one favoured placement was identified by cluster analysis. Both the

placement and flexibility of the distal C-tail were verified using site-directed fluorescence

methods applied to arrestin-1. The interaction between the distal C-tail and the N-domain is

primarily electrostatic, and salt or binding of inositol-6-phosphate disrupts this interaction. We

have further identified a functional “hinge”, which divides the relatively stable proximal C-tail

from the flexible distal C-tail. Importantly, we observe that pre-complex formation with the

phosphorylated receptor displaces the arrestin C-tail up to the hinge, and full-C-tail displacement

occurs only upon transition to the high-affinity complex. These results imply a step-by-step

displacement of the arrestin C-tail during formation of the arrestin-receptor complex.