title: The structure of the GAF A domain from phosphodiesterase 6C reveals determinants of cGMP binding, a conserved binding surface, and a large cGMP-dependent conformational change.
abstract: The photoreceptor phosphodiesterase (PDE6) regulates the intracellular levels of the second messenger cGMP in the outer segments of cone and rod photoreceptor cells. PDE6 contains two regulatory GAF domains, of which one (GAF A) binds cGMP and regulates the activity of the PDE6 holoenzyme. To increase our understanding of this allosteric regulation mechanism, we present the 2.6A crystal structure of the cGMP-bound GAF A domain of chicken cone PDE6. Nucleotide specificity appears to be provided in part by the orientation of Asn-116, which makes two hydrogen bonds to the guanine ring of cGMP but is not strictly conserved among PDE6 isoforms. The isolated PDE6C GAF A domain is monomeric and does not contain sufficient structural determinants to form a homodimer as found in full-length PDE6C. A highly conserved surface patch on GAF A indicates a potential binding site for the inhibitory subunit Pgamma. NMR studies reveal that the apo-PDE6C GAF A domain is structured but adopts a significantly altered structural state indicating a large conformational change with rearrangement of secondary structure elements upon cGMP binding. The presented crystal structure will help to define the cGMP-dependent regulation mechanism of the PDE6 holoenzyme and its inhibition through Pgamma binding.
authors.affiliation: Department of Pharmacology, University of Washington, Seattle, Washington 98195, USA.
authors.forename: Sergio E
authors.initials: SE
authors.lastname: Martinez
authors.forename: Clemens C
authors.initials: CC
authors.lastname: Heikaus
authors.forename: Rachel E
authors.initials: RE
authors.lastname: Klevit
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Beavo
mesh_terms: D000495:Allosteric Site; D000595:Amino Acid Sequence; D000818:Animals; D002645:Chickens; D018360:Crystallography, X-Ray; D006152:Cyclic GMP / Q000737:chemistry*; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6 / Q000737:chemistry*; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid
chemical_list: D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D006152:Cyclic GMP