Protein kinase A phosphorylates retinal phosducin on serine 73 in situ

Journal Article · Sat Sep 15 04:00:00 EDT 1990 · Journal of Biological Chemistry; (USA)

OSTI ID:6374365

Lee, R H; Brown, B M; Lolley, R N ^[1]

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1. Univ. of California, Los Angeles (USA)

Photoreceptors of vertebrate retinas contain a 33,000-dalton phosphoprotein, phosducin, which complexes with the beta, gamma subunits of the photoreceptor G-protein (guanine nucleotide-binding protein), transducin. In situ, the retinal content of phosphorylated phosducin is modulated by light in conjunction with light-triggered changes in intracellular cyclic nucleotide concentration. In vitro, phosducin is phosphorylated by either exogenous or endogenous protein kinase A. 32P-Labeled rat retina phosducin was isolated by immunoprecipitation either after phosphorylation by protein kinase A in the presence of (gamma-32P)ATP or after incubation of retinas in darkness with 32Pi. In either case, phosphoamino acid analysis showed that greater than 98% of 32P was linked to serine, with less than 2% to threonine. Two-dimensional peptide mapping showed that (32P)phosphoserine was associated with the same characteristic set of tryptic peptides. Furthermore, Cleveland peptide analysis using four different proteases showed that either sample exhibited identical patterns of phosphopeptides which were characteristic of the protease used. Identical phosphopeptide maps were also obtained from 32P-labeled bovine retina phosducin, indicating that the serine phosphorylation site for protein kinase A is conserved between rat and bovine. Edman degradation of phosphopeptides derived from 32P-labeled bovine phosducin showed that radioactive phosphate was incorporated into serine residue 73 which is located within a consensus phosphorylation sequence for protein kinase A (-R-K-M-S73(P)-). These observations are uniformly in agreement with protein kinase A being the endogenous kinase that phosphorylates phosducin in vivo.

• OSTI ID: 6374365
• Journal Information: Journal of Biological Chemistry; (USA), Journal Name: Journal of Biological Chemistry; (USA) Vol. 265:26; ISSN JBCHA; ISSN 0021-9258
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACID SEQUENCE
ANIMALS
ANTIBODIES
ATP
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOASSAY
BIOCHEMICAL REACTION KINETICS
BODY
BODY AREAS
CATTAILS
CATTLE
CHEMICAL REACTIONS
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DOMESTIC ANIMALS
ENZYME ACTIVITY
ENZYMES
EYES
FACE
HEAD
IMMUNOASSAY
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
LILIOPSIDA
LIQUID COLUMN CHROMATOGRAPHY
MAGNOLIOPHYTA
MAMMALS
MOLECULAR STRUCTURE
MONOCLONAL ANTIBODIES
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANS
PEPTIDES
PHOSPHORUS 32
PHOSPHORUS ISOTOPES
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHORYLATION
PHOSPHOTRANSFERASES
PLANTS
PROTEINS
RADIOISOTOPES
RATS
REACTION KINETICS
RETINA
RODENTS
RUMINANTS
SENSE ORGANS
SEPARATION PROCESSES
TRACER TECHNIQUES
TRANSFERASES
VERTEBRATES