Constitutive activation and uncoupling of the atrial natriuretic peptide receptor by mutations at the dimer interface. Role of the dimer structure in signalling

J Biol Chem. 2004 Feb 13;279(7):6115-23. doi: 10.1074/jbc.M310225200. Epub 2003 Nov 4.


The crystal packing of the extracellular hormone binding domain of the atrial natriuretic peptide (ANP) receptor contains two possible dimer pairs, the head-to-head (hh) and tail-to-tail (tt) dimer pairs associated through the membrane-distal and membrane-proximal subdomains, respectively. The tt-dimer structure has been proposed previously (van den Akker, F., Zhang, X., Miyagi, M., Huo, X., Misono, K. S., and Yee, V. C. (2000) Nature 406, 101-104). However, no direct evidence is available to identify the physiological dimer form. Here we report site-directed mutagenesis studies of residues at the two alternative dimer interfaces in the full-length receptor expressed on COS cells. The Trp74 to Arg mutation (W74R) or D71R at the hh-dimer interface caused partial constitutive guanylate cyclase activation, whereas mutation F96D or H99D caused receptor uncoupling. In contrast, mutation Y196D or L225D at the tt-interface had no such effect. His99 modification at the hh-dimer interface by ethoxyformic anhydride abolished ANP binding. These results suggest that the hh-dimer represents the physiological structure. Recently, we determined the crystal structure of ANPR complexed with ANP and proposed a hormone-induced rotation mechanism mediating transmembrane signaling (H. Ogawa, Y. Qiu, C. M. Ogata, and K. S. Misono, submitted for publication). The observed effects of mutations are consistent with the ANP-induced structural change identified from the crystal structures with and without ANP and support the proposed rotation mechanism for ANP receptor signaling.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Arginine / chemistry
  • COS Cells
  • Crystallography, X-Ray
  • Cyclic GMP / metabolism
  • DNA, Complementary / metabolism
  • Diethyl Pyrocarbonate / chemistry
  • Dimerization
  • Histidine / chemistry
  • Hydrogen Bonding
  • Kinetics
  • Mass Spectrometry
  • Models, Molecular
  • Mutagenesis, Site-Directed
  • Mutation
  • Peptides / chemistry
  • Protein Conformation
  • Receptors, Atrial Natriuretic Factor / chemistry
  • Receptors, Atrial Natriuretic Factor / genetics*
  • Signal Transduction
  • Time Factors
  • Transfection
  • Trypsin / pharmacology
  • Tryptophan / chemistry


  • DNA, Complementary
  • Peptides
  • Histidine
  • Tryptophan
  • Arginine
  • Trypsin
  • Receptors, Atrial Natriuretic Factor
  • Cyclic GMP
  • Diethyl Pyrocarbonate