A novel complex from bovine visual cells of a 33,000-dalton phosphoprotein with beta- and gamma-transducin: purification and subunit structure.
Photoreceptors of mammalian retinas contain a 33-kDa (33K) protein that is phosphorylated, in vitro, by cyclic nucleotide dependent protein kinases. The 33K protein is phosphorylated in the dark, in situ, and dephosphorylated upon illumination. The soluble 33K protein from bovine retinas has been purified to near homogeneity by extraction at pH 5.7 and chromatography on ion-exchange, gel filtration, and hydroxylapatite columns. In the native conformation, the 33K protein is associated with a 37-kDa (37K) and a 10-kDa (10K) protein, forming a trimeric complex with a sedimentation coefficient of 4.9 S and an apparent molecular mass of 77 kDa. The 33K protein can be dissociated from the 37K/10K complex by centrifugation in the presence of high pH and high salt; the subunits reassociate to form the trimeric complex upon recentrifugation in an isotonic buffer with neutral pH. The 33K protein is phosphorylated rapidly by exogenous kinase, in vitro, whereas the 37K and 10K subunits remain unphosphorylated. The 37K and 10K subunits cross-react with antibodies prepared against the beta- and gamma-subunits, respectively, of bovine transducin, indicating that the 37K and 10K subunits are immunologically identical with beta- and gamma-transducin, respectively. No immuno-cross-reactivity was observed between the 33K protein and an antibody against the alpha-subunit of bovine transducin. The 33K-beta-/gamma-transducin complex exhibits striking similarity to transducin in its subunit structure and mode of subunit interaction, suggesting it may play an important role in the metabolism and function of rod photoreceptor cells.
D000818:Animals; D001345:Autoradiography; D002417:Cattle; D002845:Chromatography / Q000379:methods; D003101:Collodion; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D005699:Galactosylceramides / Q000032:analysis / Q000138:chemical synthesis; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D010750:Phosphoproteins / Q000302:isolation & purification* / Q000378:metabolism / Q000502:physiology; D011487:Protein Conformation; D012160:Retina / Q000166:cytology*; D015293:Transducin
D005699:Galactosylceramides; D008565:Membrane Proteins; D010750:Phosphoproteins; C032840:N-palmitoylgalactosylsphingosine; D003101:Collodion; D015293:Transducin