Role of Aromaticity of Agonist Switches of Angiotensin II in the Activation of the AT1 Receptor

doi:10.1074/jbc.274.11.7103

Role of Aromaticity of Agonist Switches of Angiotensin II in the Activation of the AT₁ Receptor*

From the Department of Molecular Cardiology, Lerner Research Institute, The Cleveland Clinic Foundation, Cleveland, Ohio 44195-5069

Abstract

We have shown previously that the octapeptide angiotensin II (Ang II) activates the AT₁ receptor through an induced-fit mechanism (Noda, K., Feng, Y. H., Liu, X. P., Saad, Y., Husain, A., and Karnik, S. S. (1996)Biochemistry 35, 16435–16442). In this activation process, interactions between Tyr⁴ and Phe⁸ of Ang II with Asn¹¹¹ and His²⁵⁶ of the AT₁receptor, respectively, are essential for agonism. Here we show that aromaticity, primarily, and size, secondarily, of the Tyr⁴side chain are important in activating the receptor. Activation analysis of AT₁ receptor position 111 mutants by various Ang II position 4 analogues suggests that an amino-aromatic bonding interaction operates between the residue Asn¹¹¹ of the AT₁ receptor and Tyr⁴ of Ang II. Degree and potency of AT₁ receptor activation by Ang II can be recreated by a reciprocal exchange of aromatic and amide groups between positions 4 and 111 of Ang II and the AT₁ receptor, respectively. In several other bonding combinations, set up between Ang II position 4 analogues and receptor mutants, the gain of affinity is not accompanied by gain of function. Activation analysis of position 256 receptor mutants by Ang II position 8 analogues suggests that aromaticity of Phe⁸ and His²⁵⁶ side chains is crucial for receptor activation; however, a stacked rather than an amino-aromatic interaction appears to operate at this switch locus. Interaction between these residues, unlike the Tyr⁴:Asn¹¹¹ interaction, plays an insignificant role in ligand docking.

Footnotes

↵* This work was supported in part by National Institutes of Health Grants EY09704 and HL57470 (to S. K.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵‡ Present address: Victor Chang Cardiac Research Institute, St. Vincent’s Hospital, Darlinghurst 2010, Sydney, New South Wales, Australia.
↵§ To whom correspondence should be addressed: Dept. of Molecular Cardiology, Lerner Research Institute, The Cleveland Clinic Foundation, 9500 Euclid Ave., Cleveland, OH 44195. Tel.: 216-444-1269; Fax: 216-444-9263; E-mail: karniks{at}cesmtp.ccf.org.
Abbreviations:

Ang II

angiotensin II (DRVYIHPF)

IP

inositol phosphate

Sar

sarcosine

R

inactive receptor conformation

R′

unconstrained receptor conformation

R*

activated receptor conformation

Cha

β-cyclohexylalanine

GPCR

the G-protein-coupled receptor

DMEM

Dulbecco’s modified Eagle’s medium
- Received December 23, 1998.
The American Society for Biochemistry and Molecular Biology, Inc.