Synthetic peptides of the hamster beta 2-adrenoceptor as substrates and inhibitors of the beta-adrenoceptor kinase.
1. The beta-adrenoceptor is one of a number of G protein-coupled receptors which have been proposed to contain seven transmembrane alpha-helices. The function of this receptor appears to be regulated by phosphorylation by a specific enzyme, the beta-adrenoceptor kinase. Synthetic peptides which comprise each of the proposed intra- and extracellular domains of the beta 2-adrenoceptor have been tested as potential substrates and inhibitors of the beta-adrenoceptor kinase. 2. Two peptides which encompass the middle and terminal portions of the carboxyl tail of the receptor served as substrates by beta-adrenoceptor kinase. The kinetics of the phosphorylation reaction, however, suggest that these peptides are 10(6)-fold poorer substrate than the agonist occupied receptor. 3. A number of synthetic peptides also served as inhibitors of beta 2-adrenoceptor phosphorylation by beta-adrenoceptor kinase. In particular, a peptide which comprised the first intracellular loop of the beta 2-adrenoceptor (amino acids 56-74) inhibited most effectively with an IC50 of 40 microM. 4. These results suggest that multiple intracellular regions of the beta-receptor may serve as potential sites of interaction with beta-adrenoceptor kinase. Moreover, these regions may serve as potential targets for the development of specific inhibitors of beta-adrenoceptor kinase which could be used to block homologous desensitization.
Benovic
J L
JL
Department of Biochemistry, Howard Hughes Medical Institute, Duke University Medical Center, Durham, NC 27710.
Onorato
J
J
Lohse
M J
MJ
Dohlman
H G
HG
Staniszewski
C
C
Caron
M G
MG
Lefkowitz
R J
RJ
D000595:Amino Acid Sequence; D000818:Animals; D006224:Cricetinae; D017868:Cyclic AMP-Dependent Protein Kinases; D004591:Electrophoresis, Polyacrylamide Gel; D019204:GTP-Binding Proteins / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism* / Q000494:pharmacology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D011943:Receptors, Adrenergic, beta / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity; D051552:beta-Adrenergic Receptor Kinases
D010455:Peptides; D047428:Protein Kinase Inhibitors; D011943:Receptors, Adrenergic, beta; D011494:Protein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D019204:GTP-Binding Proteins