Role of group-conserved residues in the helical core of beta2-adrenergic receptor.
G protein-coupled receptors (GPCRs) belonging to class A contain several highly conserved (>90%) amino acids in their transmembrane helices. Results of mutational studies of these highly conserved residues suggest a common mechanism for locking GPCRs in an inactive conformation and for their subsequent activation upon ligand binding. Recently, a second set of sites in the transmembrane helices has been identified in which amino acids with small side chains, such as Gly, Ala, Ser, Thr, and Cys, are highly conserved (>90%) when considered as a group. These group-conserved residues have not been recognized as having essential structural or functional roles. To determine the role of group-conserved residues in the beta(2)-adrenergic receptor (beta(2)-AR), amino acid replacements guided by molecular modeling were carried out at key positions in transmembrane helices H2-H4. The most significant changes in receptor expression and activity were observed upon replacement of the amino acids Ser-161 and Ser-165 in H4. Substitution at these sites by larger residues lowered the expression and activity of the receptor but did not affect specific binding to the antagonist ligand dihydroalprenolol. A second site mutation, V114A, rescued the low expression of the S165V mutant. Substitution of other group-conserved residues in H2-H4 by larger amino acids lowered receptor activity in the order Ala-128, Ala-76, Ser-120, and Ala-78. Together these data provide comprehensive analysis of group-conserved residues in a class A GPCR and allow insights into the roles of these residues in GPCR structure and function.
Chelikani
Prashen
P
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Hornak
Viktor
V
Eilers
Markus
M
Reeves
Phillip J
PJ
Smith
Steven O
SO
RajBhandary
Uttam L
UL
Khorana
H Gobind
HG
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D019556:COS Cells; D002522:Chlorocebus aethiops; D017124:Conserved Sequence; D000242:Cyclic AMP / Q000378:metabolism; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000378:metabolism; D008024:Ligands; D008958:Models, Molecular; D050505:Mutant Proteins / Q000378:metabolism; D017433:Protein Structure, Secondary; D018343:Receptors, Adrenergic, beta-2 / Q000737:chemistry* / Q000378:metabolism*; D015398:Signal Transduction; D013329:Structure-Activity Relationship
D008024:Ligands; D050505:Mutant Proteins; D018343:Receptors, Adrenergic, beta-2; D000242:Cyclic AMP; D019205:GTP-Binding Protein alpha Subunits, Gs