THE PHARMACOPHORE OF THE HUMAN C5A ANAPHYLATOXIN

We have determined which amino acids contribute to the pharmacophore o f human C5a, a potent inflammatory mediator. A systematic mutational a nalysis of this 74-amino acid protein was performed and the effects on the potency of receptor binding and of C5a-induced intracellular calc ium ion mobilization were measured. This analysis included the constru ction of hybrids between C5a and the homologous but unreactive C3a pro tein and site-directed mutagenesis. Ten noncontiguous amino acids from the structurally well-defined 4-helix core domain (amino acids 1-63) and the C-terminal arginine-containing tripeptide were found to contri bute to the pharmacophore of human C5a. The 10 mostly charged amino ac ids from the core domain generally made small incremental contribution s toward binding affinity, some of which were independent. Substitutio ns of the C-terminal amino acid Arg 74 produced the largest single eff ect. We also found the connection between these 2 important regions to be unconstrained.