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 1 , Ciara C.M. Lally 1 , Brian Bauer 1 , Jana Selent 2 . 2 Pompeu Fabra University, Hospital del Mar Medical Research Institute, Barcelona, Spain. 1 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.

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