Synthesis of a new photoreactive C5a analog that permits identification of the ligand binding component of the granulocyte C5a receptor.

Human C5a is a complement derived inflammatory mediator that binds to specific receptors that are found in the granulocyte plasma membrane. A new photoreactive C5a analog, that can be radiolabeled in the aryl azide moiety, has been synthesized by coupling p-azidosalicyl-2-mercapto-N-ethylamide-2'-thiopyridine disulfide to a partially reduced form of C5a. This unique analog (ASAMEA-SC5a) specifically bound to the granulocyte C5a receptor with an apparent Kd of 1.8 nM, a value that is comparable to that of the native ligand. Photoactivation resulted in crosslinking of this probe to the granulocyte C5a receptor. Subsequent reductions of the complex produced a radiolabeled C5a receptor, or a subunit of this receptor that had an apparent molecular weight of 47,000 Daltons on SDS-PAGE.