In crustaceans, molting hormone (ecdysteroid) induces molting like in insects. In the American crayfish (Procambarus clarkii) during the process toward molting various events occur; most calcium carbonate in the cuticle is dissolved (decalcification) and transferred to the stomach to become a pair of gastroliths. After molting, the gastroliths are dissolved again and transferred to newly formed cuticle to harden the exoskeleton. An acid-insoluble, SDS/DTT-soluble protein named gastrolith matrix protein (GAMP) was purified from gastroliths of the American crayfish. Based on the partial amino acid sequences determined, a cDNA encoding GAMP was cloned. GAMP consists of 462 amino acid residues with two kinds of characteristic repeated sequences. GAMP showed calcification inhibitory activity in vitro. By immunohistochemistry, GAMP was found to be present in the gastrolith-producing epithelial cells (gastrolith discs) in the stomach and gastroliths themselves. Gastroliths and exoskeleton contain amorphous calcium carbonate (ACC), which was found to be stabilized by glycolytic intermediates, phosphoenolpyruvate, and 3-phosphoglycerate.