Role of Phe308 in the seventh transmembrane domain of the AT2 receptor in ligand binding and signaling

• Published in: Biochemical and biophysical research communications Vol. 319; no. 4; pp. 1138 - 1143
• Main Authors: Pulakat, Lakshmi, Mandavia, Chirag H, Gavini, Nara
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 09.07.2004
• Subjects: Amino Acid Sequence; Angiotensin II; Angiotensin II - metabolism; Animals; AT1 receptor; AT2 receptor; Cyclic CMP - metabolism; Humans; Ligands; M8 mutant of the AT2; Molecular Sequence Data; Mutagenesis, Site-Directed; Oocytes - physiology; Phe308Ala; Phenylalanine - metabolism; Protein Binding; Protein Structure, Secondary; Protein Structure, Tertiary; Rats; Receptor, Angiotensin, Type 2 - chemistry; Receptor, Angiotensin, Type 2 - genetics; Receptor, Angiotensin, Type 2 - metabolism; Signal Transduction - physiology; Tyrosine - metabolism; Xenopus; Xenopus oocytes; Xenopus oocytes; Phe308Ala; Angiotensin II; AT2 receptor; M8 mutant of the AT2; AT1 receptor
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2004.05.092

Studies on Angiotensin II (Ang II) receptor type AT1 have suggested that interaction between the two highly conserved residues, Tyr292 in the 7th transmembrane domain (TMD) and the Asp74 in the 2nd TMD, is critical for linking the Ang II binding and AT1 receptor-Gq protein coupling. In the Ang II receptor type AT2, the Asp is conserved (Asp90 in 2nd TMD), however, there is no Tyr residue in the 7th TMD and Phe308 occupies the analogous position to Tyr292 of the AT1. Replacing this Phe308 with Ala reduced receptor affinity to peptidic ligands 125I-Ang II ( K d=0.37 nM) and 125I-CGP42112A ( K d=0.56 nM), but retained the ability of the AT2 to reduce cGMP levels in Xenopus oocytes. Thus, the Phe308 of the AT2 does not mimic the role of Tyr292 of the AT1 in the receptor activation upon Ang II binding. We have also shown that the M8 mutant of the AT2 with the 7th TMD similar to that of wild type AT2 can couple to PLC like the AT1 and bind the AT2-specific ligands with high affinity [FEBS Lett. 532 (2002) 379]. Since the Ang II is shown to bind to both the AT1 and the AT2 in an identical manner, we propose that the absence of Tyr in the 7th TMD of the AT2 does not prevent the receptor from coupling to Gq-protein, rather may contribute to the freedom of the AT2 to couple to trimeric G-proteins in both G-βγ dependent and independent manners upon Ang II binding.