A novel rhodopsin kinase in octopus photoreceptor possesses a pleckstrin homology domain and is activated by G protein betagamma-subunits.
G protein-coupled receptor kinases (GRKs) play an important role in stimulus-dependent receptor phosphorylation and desensitization of the receptors. Mammalian rhodopsin kinase (RK) and beta-adrenergic receptor kinase (betaARK) are the most studied members among known GRKs. In this work, we purified RK from octopus photoreceptors for the first time from invertebrate tissues. The molecular mass of the purified enzyme was 80 kDa as estimated by SDS-polyacrylamide gel electrophoresis, and this was 17 kDa larger than that of the vertebrate enzymes. Unlike vertebrate RK, octopus RK (ORK) was directly activated by betagamma-subunits of a photoreceptor G protein. We examined the effects of various known activators and inhibitors of GRKs on the activity of the purified ORK and found that their effects were different from those on either bovine RK or betaARK. To analyze the primary structure of the enzyme, we cloned the cDNA encoding ORK from an octopus retinal cDNA library. The deduced amino acid sequence of the cDNA was highly homologous to betaARK over the entire molecule, including a pleckstrin homology domain located in the C-terminal region, and homology to RK was significantly lower. Furthermore, Western blot analysis of various octopus tissues with an antibody against the purified ORK showed that ORK is expressed solely in the retina, which confirmed the identity of the enzyme as rhodopsin kinase. Thus, ORK appears to represent a unique subgroup in the GRK family, which is distinguished from vertebrate RK.
Kikkawa
S
S
Department of Life Science, Himeji Institute of Technology, Harima Science Garden City, Akoh-gun, Hyogo 678-1279, Japan.
Yoshida
N
N
Nakagawa
M
M
Iwasa
T
T
Tsuda
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001798:Blood Proteins / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D004789:Enzyme Activation; D005136:Eye Proteins / Q000737:chemistry / Q000378:metabolism*; D051606:G-Protein-Coupled Receptor Kinase 1; D019204:GTP-Binding Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D049831:Octopodiformes; D010750:Phosphoproteins; D010766:Phosphorylation; D017956:Photoreceptor Cells, Invertebrate / Q000737:chemistry / Q000378:metabolism*; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism* / Q000493:pharmacokinetics; D016415:Sequence Alignment; D014018:Tissue Distribution
D001798:Blood Proteins; D005136:Eye Proteins; D010750:Phosphoproteins; C039077:platelet protein P47; D011494:Protein Kinases; D051606:G-Protein-Coupled Receptor Kinase 1; D019204:GTP-Binding Proteins