The inhibitory gamma subunit of the rod cGMP phosphodiesterase binds the catalytic subunits in an extended linear structure.
The unique feature of rod photoreceptor cGMP phosphodiesterase (PDE6) is the presence of inhibitory subunits (Pgamma), which interact with the catalytic heterodimer (Palphabeta) to regulate its activity. This uniqueness results in an extremely high sensitivity and sophisticated modulations of rod visual signaling where the Pgamma/Palphabeta interactions play a critical role. The quaternary organization of the alphabetagammagamma heterotetramer is poorly understood and contradictory patterns of interaction have been previously suggested. Here we provide evidence that supports a specific interaction, by systematically and differentially analyzing the Pgamma-binding regions on Palpha and Pbeta through photolabel transfer from various Pgamma positions throughout the entire molecule. The Pgamma N-terminal Val16-Phe30 region was found to interact with the Palphabeta GAFa domain, whereas its C terminus (Phe73-Ile87) interacted with the Palphabeta catalytic domain. The interactions of Pgamma with these two domains were bridged by its central Ser40-Phe50 region through interactions with GAFb and the linker between GAFb and the catalytic domain, indicating a linear and extended interaction between Pgamma and Palphabeta. Furthermore, a photocross-linked product alphabetagamma(gamma) was specifically generated by the double derivatized Pgamma, in which one photoprobe was located in the polycationic region and the other in the C terminus. Taken together the evidence supports the conclusion that each Pgamma molecule binds Palphabeta in an extended linear interaction and may even interact with both Palpha and Pbeta simultaneously.
Guo
Lian-Wang
LW
Department of Pharmacology, University of Wisconsin Medical School, Madison, Wisconsin 53706, USA. lianwangguo@wisc.edu
Muradov
Hakim
H
Hajipour
Abdol R
AR
Sievert
Michael K
MK
Artemyev
Nikolai O
NO
Ruoho
Arnold E
AE
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain; D002417:Cattle; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D005136:Eye Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D066298:In Vitro Techniques; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010777:Photochemistry; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D017948:Retinal Rod Photoreceptor Cells / Q000201:enzymology*
D005136:Eye Proteins; D021122:Protein Subunits; D010727:Phosphoric Diester Hydrolases; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6