CLONING AND CHARACTERIZATION OF THE GENE ENCODING AN EXTRACELLULAR ALKALINE SERINE-PROTEASE FROM VIBRIO-METSCHNIKOVII STRAIN-RH530

The gene vapT, encoding VapT, one of the extracellular sodium dodecyl sulfate (SDS)-resistant alkaline serine proteases (Serp) from the Gram (-) Vibrio metschinikovii strain RH530 has been cloned in Escherichia coli. The recombinant E. coli produced a protease which co-migrated wi th VapT on gelatin polyacrylamide gels. The nucleotide (nt) sequence o f the cloned vapT revealed a single open reading frame of 1641 bp enco ding 547 amino acids (aa) (58 961 Da). Upon analysis of the N-terminal aa sequence, VapT was shown to be processed properly in recombinant E . coli and to consist of 428 aa (45 626 Da). The deduced aa sequence o f VapT showed significant sequence homology to subtilisin Carlsberg fr om Bacillus licheniformis, particularly in the regions containing acti ve site residues and calcium-binding sites. VapT had an intervening re gion of approx. 149 aa between the His and Ser residues of the active site, as compared with other Serp.