Proteinase inhibitor from Stichodactyla helianthus: purification, characterization and immobilization

A Kunitz type proteinase inhibitor was isolated from extracts of the sea anemone Stichodactyla helianthus. The purification procedure comprises treatment with trichloroacetic acid followed by affinity chromatography on trypsin-Sepharose and gel filtration on Sephadex G-50 or ion exchange chromatography on CM-cellulose. The major inhibitor (isoelectric point 8.4) is a small protein consisting of 55 amino acid residues lacking tryptophan and methionine, with a molecular weight of 6110 (FAB-MS). The amino acid sequence and the structure in solution (NMR) were determined. The inhibitor exhibits a broad specificity for serine, cysteine and aspartic proteinases. Reactors were prepared with immobilized inhibitor in different supports (Sepharose, cellulose, and silica gel) to be employed in the elimination of undesirable proteinases and the purification of different kinds of proteolytic enzymes.