Cloning and characterization of the gene encoding an extracellular alkaline serine protease from Vibrio metschnikovii strain RH530.
The gene vapF, encoding VapT, one of the extracellular sodium dodecyl sulfate (SDS)-resistant alkaline serine proteases (Serp) from the Gram- Vibrio metschnikovii strain RH530 has been cloned in Escherichia coli. The recombinant E. coli produced a protease which co-migrated with VapT on gelatin polyacrylamide gels. The nucleotide (nt) sequence of the cloned vapT revealed a single open reading frame of 1641 bp encoding 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 of VapT showed significant sequence homology to subtilisin Carlsberg from Bacillus licheniformis, particularly in the regions containing active site residues and calcium-binding sites. VapT had an intervening region of approx. 149 aa between the His and Ser residues of the active site, as compared with other Serp.
Kwon
Y T
YT
Department of Molecular Biology, Seoul National University, South Korea.
Kim
J O
JO
Moon
S Y
SY
Yoo
Y D
YD
Rho
H M
HM
D000468:Alkalies; D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism; D017422:Sequence Analysis, DNA; D012697:Serine Endopeptidases / Q000235:genetics*; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013379:Substrate Specificity; D014508:Urea / Q000494:pharmacology; D014733:Vibrio / Q000201:enzymology / Q000235:genetics*
D000468:Alkalies; D001426:Bacterial Proteins; D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate; D014508:Urea; D012697:Serine Endopeptidases; C091638:VapT protein, Vibrio metschnikovii; D002118:Calcium