IDENTIFICATION AND CHARACTERIZATION OF AN EXTRACELLULAR PROTEASE ACTIVITY PRODUCED BY THE MARINE VIBRIO SP-60

The marine fish pathogen Vibrio sp. 60 has been used as a host for het erologous expression of the Escherichia coli heat-labile enterotoxin B -subunit and derivatives carrying a C-terminal extension. In this stud y, a chimeric enterotoxin B-subunit with an extension corresponding to the carboxy-terminal nine amino acids -Tyr-Ala-Gly-Ala-Val-Val-Asn-As p-Leu-COOH from the small subunit of herpes simplex virus type 1-encod ed ribonucleotide reductase, is shown to be proteolytically cleaved in the extracellular medium by a single protease that is secreted by the host strain. Such protease behaves as a typical metalloprotease, bein g inhibited by EDTA but not by a serine protease inhibitor. Purificati on and amino acid composition analysis of the two proteolysis products revealed a specific cleavage of the peptide-bond between amino acids glycine and alanine of the nine amino acid extension with loss of acti vity. The above observation is relevant for the biotechnological explo itation of Vibrio sp. 60.