Three-dimensional structure of non-activated cGMP phosphodiesterase 6 and comparison of its image with those of activated forms.
Cyclic GMP phosphodiesterase (PDE6) in rod photoreceptors, a key enzyme in vertebrate phototransduction, consists of two homologous catalytic subunits (Palpha and Pbeta) and two identical regulatory subunits (Pgammas). Pgamma regulates the PDE activity through its direct interaction with transducin. Here, using electron microscopy and image analysis of single particles, we show the three-dimensional organization of the basic form of bovine PDE, Palphabetagammagamma, and compare its average image with those of Pgamma-released PDE. The structure of Palphabetagammagamma appears to be a flattened bell-shape, with dimensions of 150 x 108 x 60A, and with a handle-like protrusion attached to the top of the structure. Except for the protrusion, the organization consists of two homologous structures arranged side by side, with each structure having three distinct regions, showing pseudo twofold symmetry. These characteristics are consistent with a model in which the overall structure of Palphabetagammagamma is determined by hetero-dimerization of Palpha and Pbeta, with each subunit consisting of one catalytic and two GAF regions. A comparison of the average image of Palphabetagammagamma with those of Pgamma-released PDE suggests that Pgamma release does not affect the overall structure of Palphabeta, and that the Palphabeta C-terminus, but not Pgamma, is a determinant for the Palphabeta orientation on carbon-coated grids. These observations suggest that the basic structure of PDE does not change during its regulation, which implies that Palphabeta is regulated by its regional interaction with Pgamma.
Kajimura
Naoko
N
Biomolecular Engineering Research Institute, 6-2-3, Furuedai, Suita, Osaka 565-0874, Japan.
Yamazaki
Matsuyo
M
Morikawa
Kosuke
K
Yamazaki
Akio
A
Mayanagi
Kouta
K
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000737:chemistry*; D000818:Animals; D002417:Cattle; D018360:Crystallography, X-Ray; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D019281:Dimerization; D005136:Eye Proteins; D007091:Image Processing, Computer-Assisted; D008854:Microscopy, Electron; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D017948:Retinal Rod Photoreceptor Cells / Q000201:enzymology / Q000648:ultrastructure
D005136:Eye Proteins; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6