The IQ domains in neuromodulin and PEP19 represent two major functional classes.

The affinities of Ca(2+)-saturated and Ca(2+)-free calmodulin for a fluorescent reporter construct containing the PEP19 IQ domain differ by a factor of approximately 100, with K(d) values of 11.0 +/- 1.2 and 1128.4 +/- 176.5 muM, respectively, while the affinities of a reporter containing the neuromodulin IQ domain are essentially identical, with K(d) values of 2.9 +/- 0.3 and 2.4 +/- 0.3 muM, respectively. When Ca(2+) is bound only to the C-terminal pair of Ca(2+)-binding sites in calmodulin, the K(d) value for the PEP19 reporter complex is decreased approximately 5-fold, while the value for the neuromodulin reporter complex is increased by the same factor. When Ca(2+) is bound only to the N-terminal pair of Ca(2+)-binding sites, the K(d) value for the PEP19 reporter complex is unaffected, but the value for the complex with the neuromodulin reporter is increased approximately 12-fold. These functional differences are largely ascribed to three differences in the CaM-binding sequences of the two reporters. Replacement of a central Gly in the neuromodulin IQ domain with a Lys at this position in PEP19 almost entirely accounts for the distinctive patterns of Ca(2+)-dependent stability changes exhibited by the two complexes. Replacement of a Lys immediately before the "IQ" amino acid pair in the neuromodulin sequence with the Ala in PEP19 accounts for the remaining Ca(2+)-dependent differences. Replacement of an Ala in the N-terminal half of the neuromodulin sequence with the Gln in PEP19 accounts for approximately half of the Ca(2+)-independent difference in the stabilities of the two reporter complexes, with the Ca(2+)-independent effect of the Lys replacement accounting for most of the remainder. Since the central Gly in the neuromodulin sequence is conserved in half of all known IQ domains, these results suggest that the presence or absence of this residue defines two major functional classes.