ABSTRACT
Members of the cGMP phosphodiesterase (PDE6) family are the key photoreceptor proteins regulating the concentration of the second messenger, cGMP, in response to light in cones and rods. Rod PDE6 consists of two catalytic subunits PDE6α and PDE6β, which form a heterodimer. The cone-specific PDE6, PDE6α′, forms a homodimer in cones. Under dark conditions, both PDE6s are inhibited by two copies of rod-and cone-specific subunits, Pγ, respectively. The domain structure of the catalytic subunits of PDE6 is very similar to another member of the PDE family, PDE5 (1-4). However, the unique functional features of PDE6 that distinguish it from PDE5 and all other members of the PDE family are the presence of the tightly bound inhibitory Pγ subunits and a very high kcat value approaching the diffusion barrier. The inhibition is relieved by interaction of the Pγ subunits with transducin-α (Tα)-GTP to allow robust PDE activation and rapid decrease in the concentration of cGMP, which in turn leads to the closure of cGMP-gated Na/Ca channels and hyperpolarization of the photoreceptor cells. Here, we describe procedures routinely used in our laboratory to study the function and regulation of PDE6.
