PALMITOYLATION OF THE ALPHA(2A)-ADRENERGIC RECEPTOR - ANALYSIS OF THESEQUENCE REQUIREMENTS FOR AND THE DYNAMIC PROPERTIES OF ALPHA(2A)-ADRENERGIC RECEPTOR PALMITOYLATION

Previous studies have demonstrated that the alpha(2A)-adrenergic recep tor (alpha(2A)AR) incorporates [H-3]palmitate and that replacement of Cys(442) by Ala or Ser eliminates detectable acylation without perturb ing coupling to pertussis toxin-sensitive GTP-binding proteins (Kenned y, M. E., and Limbird, L. E. (1993) J. Biol. Chem. 268, 8003-8011) or, as shown here, without perturbing agonist-dependent receptor phosphor ylation, in contrast to the consequences of eliminating beta(2)-adrene rgic receptor acylation, As a first step in revealing the functional r ole for this post-translational modification at the alpha(2A)AR, we ex plored sequences in the alpha(2A)AR which confer alpha(2A)AR acylation and whether or not [H-3]palmitoylation of the alpha(2A)AR is dynamic, Deletion of the 7 terminal amino acids distal to Cys(442) Of the alph a(2A)AR did not eliminate detectable [H-3]palmitoylation of the alpha( 2A)AR, whereas truncation to Leu(441) did, indicating both that Cys(44 2) is the likely site for acylation and that sequences distal to Cys(4 42) are not required for acylation at Cys(442), Since mutation of sequ ences proximal to Cys(442) altered overall receptor structure, based o n markedly reduced detectable adrenergic receptor binding, proximal mo tifs required for palmitoylation of the alpha(2A)AR could not be explo red fur ther. When the turnover of [S-35]Met/Cys-labeled alpha(2A)AR w as compared with the turnover of the [H-3]palmitate-labeled alpha(2A)A R, it was of interest that agonist treatment accelerated the half-life of decay of the [H-3]palmitate-labeled alpha(2A)AR without detectable receptor down-regulation, providing evidence that the acylation of th e alpha(2A)AR may be a dynamic process.