HATCHING ENZYME FROM THE SEA-SQUIRT CIONA-INTESTINALIS - PURIFICATIONAND PROPERTIES

We have purified a 34 kDa hatching enzyme from the water in which the embryos of the sea-squirt Ciona intestinalis hatch. This enzyme was ob tained in homogeneous form as judged from SDS-PACE and HPLC gel filtra tion. The enzyme possesses proteolytic activity and is able to digest the chorion of the egg of C. intestinalis. It is a metalloproteinase a nd contains one atom of Zn per molecule. The optimum pH is 8.5. The en zyme shows hydrolytic activity towards the -CO-NH- bonds, which are hy drolyzed by the members of the serine proteinase family. It has a tryp sin-like activity in that it cuts the bond of Arg and Lys at P-1 posit ion of the scissile bond -P-1-P-1', but it differs from trypsin insofa r as it hydrolyzes the peptide bond on either side of Arg and Lys. The purified enzyme is inhibited by the common metal-chelators and by the classical trypsin proteinase inhibitors. The apparent K-m, values at 37 degrees C and pH 8.5 toward tosyl-Gly-Pro-Arg-NHNap, tosyl-Gly-Pro- Lys-NHNap and Bz-Arg-Gly-Arg-NHNap were 0.125, 0.5 and 2.5 mM, respect ively. The results obtained in this study suggest that the hatching en zyme from C. intestinalis exhibits both trypsin-like activity and meta lloproteinase activity.