Molecular analysis of an extracellular protease gene from Vibrio parahaemolyticus.
The structural gene prtVp encoding the extracellular protease of Vibrio parahaemolyticus strain 93 was cloned in Escherichia coli and sequenced. The cloned DNA fragment contained a 1761 bp ORF encoding a 587 amino acid protein. The deduced polypeptide is composed of a 25 amino acid signal peptide and a 562 amino acid extracellular polypeptide with a calculated molecular mass of 63,156 Da. Protease analysis using a gelatin-containing SDS-polyacrylamide gel detected the presence of a 62 kDa protease that was present in the culture supernatant fractions of the wild-type V. parahaemolyticus strain and of E. coli bearing a pUC119 recombinant with the prtVp DNA insert. The protease activity was inhibited by zinc- and metal-specific inhibitors such as EDTA and 1,10-phenanthroline, which suggested that it is a metalloprotease. The deduced amino acid sequence of PrtVp has 32% identity with that of the collagenase of Vibrio alginolyticus, but has no identity with those of the bacterial proteases. A conserved zinc-binding domain was also found in PrtVp from homology comparison with other metalloproteases. This PrtVp can cause weak haemolysis on blood agar.
Lee
C Y
CY
Graduate Institute of Agricultural Chemistry, National Taiwan University, Taipei, Republic of China.
Su
S C
SC
Liaw
R B
RB
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D005798:Genes, Bacterial; D006461:Hemolysis; D008666:Metalloendopeptidases / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011480:Protease Inhibitors / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis; D017386:Sequence Homology, Amino Acid; D014733:Vibrio; D014736:Vibrio parahaemolyticus; D015032:Zinc / Q000378:metabolism
D001426:Bacterial Proteins; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D008666:Metalloendopeptidases; C073584:Prt protein, Streptomyces; D015032:Zinc