Purification and Characterization of an Elastolytic Protease of Vibrio vulnificus

SUMMARY: Large amounts of a highly purified, extracellular elastolytic protease of Vibrio vulnificus were obtained by sequential ammonium sulphate precipitation and hydrophobic interaction chromatography with phenyl-Sepharose CL-4B. The protease had an M r of about 50500 (estimated by SDS-PAGE), a pI of 5·7, and a temperature optimum range of 55 to 60 °C. The pH optimum and the results of inactivation studies suggested that the enzyme was a neutral metalloprotease. The protease had about 429 amino acid residues, and the first 20 amino-terminal amino acid residues were Ala-Gln-Ala-Asn-Gly-Thr-Gly-Pro-Gly-Gly-Asn-Ser-Lys-Thr-Gly-Arg-Tyr-Glu-Phe-Gly. The purified protease was toxic for mice (about 1·5 mg kg−1and 4·5 mg kg−1, intraperitoneal and intravenous LD50 values, respectively), and subcutaneous injection of the enzyme elicited rapid and extensive dermonecrosis.