PT - JOURNAL ARTICLE AU - Clayton M. Bullock AU - Jia-Da Li AU - Qun-Yong Zhou TI - Structural Determinants Required for the Bioactivities of Prokineticins and Identification of Prokineticin Receptor Antagonists AID - 10.1124/mol.65.3.582 DP - 2004 Mar 01 TA - Molecular Pharmacology PG - 582--588 VI - 65 IP - 3 4099 - http://molpharm.aspetjournals.org/content/65/3/582.short 4100 - http://molpharm.aspetjournals.org/content/65/3/582.full SO - Mol Pharmacol2004 Mar 01; 65 AB - Prokineticins are cysteine-rich secreted proteins that regulate diverse biological processes, including gastrointestinal motility, angiogenesis, and circadian rhythms. Two closely related G protein-coupled receptors that mediate signal transduction of prokineticins have recently been cloned. The structural elements required for prokineticins' bioactivities are still unknown. We show here that both the N-terminal hexapeptide (AVITGA) and C-terminal cysteine-rich domains are critical for the bioactivities of prokineticins. Substitutions, deletions, and insertions to the conserved N-terminal hexapeptides result in the loss of agonist activity. Mutant prokineticins with the substitution of the first N-terminal alanine with methionine or the addition of a methionine to the N terminus inhibit the activation of prokineticin receptors and thus are considered as antagonists of prokineticin receptors. We have further shown that mutations in selected cysteine residues in the C-terminal domain result in prokineticins without biological activity. The essential role of C-terminal domain is reinforced by two observations: that peptides without the carboxyl domain and proteins with the N-terminal hexapeptide fused to the carboxyl domains of colipase or dickkopf are devoid of biological activity. We demonstrate that limited structural changes of C-terminal cysteine-rich regions of prokineticins are tolerable because chimeric prokineticins with swapped cysteine-rich domains between prokineticin 1 and prokineticin 2, as well as a splice variant of prokineticin 2 that contains extra 21 residue insertion in its C-terminal domain, are biologically active.