Coupling an alpha 2-adrenergic receptor peptide to G-protein: a new photolabeling agent

A photoreactive derivative of a tetradecapeptide G-protein activator (peptide Q) derived from the alpha 2-adrenergic receptor was designed and used to label purified G-protein (Go/Gi). N-bromoacetyl-N'-(3-diazopyruvoyl)-m-phenylene-diamine (Br-DAP) was conjugated to the C-terminal cysteine of peptide Q. The DAP-modified peptide Q (DAP-Q) specifically incorporated into the a subunit of Go. The incorporation of DAP-Q into alpha o was blocked by unmodified Q peptide (IC50 = 15 +/- 6 microM; n = 4). Photolysis of sixfold higher concentrations of DAP-Q with ovalbumin or bovine albumin failed to produce cross-linked products. Br-DAP should prove useful in detecting mutual contact sites between peptides and their binding proteins.