Biomedical Engineering Reference
In-Depth Information
The 7-TMH domain (7TMD) is the most structurally conserved region of
GPCRs although its sequence conservation is only moderate, with sequence identi-
ties for 7TMDs ranging from about 70 % for receptors of the same subfamily (e.g.,
the opioid receptor subfamily, the adrenergic receptor subfamily), to a mere 17 %
(well within the homology twilight zone) between the protease activated receptor 1
(PDB code 3vw7) and the sphingosine 1-phosphate receptor (PDB code 2v2y), see
Table
3.2
. In spite of this relatively low sequence identity between families, a num-
ber of highly conserved residues and motifs are found in the transmembrane helices.
The most conserved residue in each TMH forms the basis of the Ballesteros-Wein-
stein numbering system [
33
] in which this residue gets the number 50, residues to
the N-terminus of this residue get numbers smaller than 50 and residues to its C-
terminus get numbers greater than 50. Each residue within the 7TMD is thus identi-
fied by two numbers: the TMH number and a number indicating its relative position
with respect to the most conserved one in this TMH. For instance, 7.49 represents
the residue preceding the most conserved one (a Proline) in the TMH7 sequence.
Ballesteros-Weinstein 50 (N50) residues are highlighted in yellow in Fig.
3.2
. Func-
tionally important sequence motifs for GPCRs are also found in the TMHs (shown
in bold in Fig.
3.2
), for instance the D[E]RY motif at the end of TMH3 that is part
of a “ionic lock”, the FxxCWxP motif in TMH6 and NPxxY motif at the end of
TMH7 that are suspected to be involved in the activation mechanisms (see next sec-
tion). The presence of these conserved residues facilitates the alignment of GPCR
sequences. However, N50 residues for TMH5 and TMH6 are not conserved in ol-
factory receptor sequences. In particular, obtaining an accurate alignment for OR
TMH6 is challenging since the CWxP motif is not preserved in most OR sequences.
Loops are more diverse in terms of length, sequence similarity and 3D confor-
mation. There exist two types of structures, those in which the binding pocket is
occluded by the N-terminus and the ECLs (e.g., rhodopsin and sphingosine 1-phos-
phate receptors) and those in which the binding pocket is accessible from the extra-
cellular side. ECL2 exhibits a more pronounced structural diversity than ECL1 and
ECL3 that are short loops, respectively 5-6 and 7-8 residue long. In most GPCRs,
ECL2 that connects TMH4 to TMH5 is tethered to TMH3 by a disulfide bridge
between Cys
3.25
at the extracellular tip of TMH3 and a highly conserved Cys in the
loop. This generates two “pseudo-loops”: ECL2a and ECL2b. In a number of struc-
tures, ECL2b acts as a lid on top of the binding pocket and its residues interact with
the ligand. The conformation of the 6-residue long ICL1 is conserved in all known
structures. The 9-12 residue long ICL2 that has been shown to interact with the N-
terminus of the Gα subunit [
30
] is very flexible and adopts diverse conformations
in the known structures. ICL3 has a highly variable length, ranging from 5 residues
in CXCR4 up to tens of residues in some other receptors. Presumably, ICL3 is
responsible of the receptor affinity for different G proteins.
The ligand binding pocket is located in the first one-third of the helical bundle
from the extracellular side. GPCRs bind very different ligands (proteins, peptides,
lipids, nucleotides, small organic molecules, ions) that exhibit diverse shapes,
sizes and chemical properties and penetrate to different depths in the pocket. Ven-
katakrishnan and coll. have analyzed the positions in the binding pocket at which
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