Equistatin, a protease inhibitor from the sea anemone Actinia equina, is composed of three structural and functional domains

A cDNA encoding a precursor of equistatin, a potent cysteine and aspartic p roteinase inhibitor, was isolated from the sea anemone Actinia equina. The deduced amino acid sequence of a 199-amino-acid residue mature protein with 20 cysteine residues, forming three structurally similar thyroglobulin typ e-1 domains, is preceded by a typical eukaryotic signal peptide. The mature protein region and those coding for each of the domains were expressed in the periplasmic space of Escherichia coli, isolated, and characterized. The whole recombinant equistatin and its first domain, but not the second and third domains, inhibited the cysteine proteinase papain (K-i 0.60 nM) compa rably to natural equistatin. Preliminary results on inhibition of cathepsin D, supported by structural comparison, show that the second domain is like ly to be involved in activity against aspartic proteinases. (C) 2000 Academ ic Press.