ER-Bound Steps in the Biosynthesis of G Protein-Coupled Receptors - GPCR Signalling Complexes: Synthesis, Assembly, Trafficking and Specificity - page 12

export-de fi cient V 2 -vasopressin receptor into an arti fi cial single-pass transmembrane
protein (assembled from the N-terminus, transmembrane helix I and the receptor
C-terminus of the V 2 -receptor) failed to suppress export to the cell surface (Thielen
et al. 2005 ). The export signal therefore may be functional only in the context
of the entire receptor polypeptide.
Alternatively, the presence of a native helix VIII structure may serve as an
indication of the correct fold, which should allow for clearance by the ER quality
control (see below). It is reasonable to predict that folding of helix VIII is a
thermodynamically favorable event. Molecular dynamics simulation on the CB1
receptor suggests that helix VIII buttresses a hydrophobic pocket lined by neigh-
boring side chains from transmembrane helices I and VII and CL-1. The pocket is
so hydrophobic that it attracts the hydrophobic acyl chains of the surrounding
lipids. The agglomeration of protein and lipids (including the palmitoyl cystein-
thioester that borders helix VIII) goes along with a marked decline in free energy
and the resulting hydrophobic pocket contributes to energy minimization of the
global receptor structure (Shim 2009 ). Thus, helix VIII folds spontaneously after
membrane insertion of the receptor polypeptide and - in a bolt-like fashion - closes
the circular configuration.
The efficacy of pharmacochaperones varies depending on the state of the proxi-
mal C-tail. In the A 1 adenosine receptor exchange of a conserved helix VIII
phenylalanine for alanine causes ER retention of the mutant receptor that is
rescued by ligand occupancy. By contrast, complete truncation of the membrane
proximal C-terminus produces a receptor refractory to pharmacochaperone rescue
(Pankevych et al. 2003 ; Hawtin 2006 ; Málaga-Diéguez et al. 2010 ) . Ligand
occupancy causes a conformational change in the transmembrane domain and may
force packing of the helix VIII pocket via the interhelical network of side-chain
and backbone bonds.

1.5.1 ER Quality Control

We presume that the arrangement of the transmembrane a-helices, helix VIII and
the extracellular disulfide bonds represent minimum structural criteria of G protein
coupled receptors that have to be met before receptors are released from the ER.
Compared to this well-arranged core and in the absence of interaction partners,
cytoplasmic loops 2 and 3 and the distal c-tail are highly flexible. Similarly, the
N-terminal tail of many rhodopsin related receptors (as opposed to receptors of the
glutamate, secretin, adhesion and frizzled receptor subfamilies) does not adopt a
stable conformation. Therefore, much of the cytoplasmic domain and of the N tail
must be less important for clearance by quality control than that of the stable core
structure.
The vast majority of cargo present in the ER including receptors exposes unfolded
or misfolded determinants that elicit chaperone binding. Chaperone molecules
involved in quality control have been originally described to operate in the ER-lumen