Photoinactivation and identification of the biogenic amine transporter in chromaffin granules from bovine adrenal medulla

We have synthesized a new derivative of 5-hydroxytryptamine, a substrate of the biogenic amine transporter in bovine chromaffin granules. This new compound, 4-azido, 3-nitrophenyl-azo-(5-hyroxytryptamine), is a competitive, reversible, and specific inhibitor of the biogenic amine transporter in the dark (K(i) = 2 μM). Upon illumination, the inactivation of the transporter becomes irreversible and follows an apparent first order rate kinetics (K(INAC) = 4 μM). The transporter can be protected if other substrates are present during illumination. When [³H]4-azido,3-nitrophenyl-azo-(5-hydroxytryptamine) is used, preferential labeling of one polypeptide (M(r) ~ 45,000) is observed in the light, provided no other substrates of the transporter are present during illumination. We have tentatively identified this polypeptide as the transporter itself or one of its subunits.