Use of a gram- signal peptide for protein secretion by gram+ hosts: basic protease of Dichelobacter nodosus is produced and secreted by Bacillus subtilis.
The bprV gene, encoding the extracellular basic protease of the Gram- anaerobic bacterium Dichelobacter nodosus, was expressed and the protein secreted in Bacillus subtilis using the novel cloning/expression vector pNC3 [Wu et al., Gene 106 (1991) 103-107]. The pre- and pro-peptides were processed correctly in this heterologous system, and the 127-amino acid C-terminal extension region was also removed. The recombinant gene product was indistinguishable biochemically or immunochemically from the authentic protease and was able to form crystals upon dialysis, as was found for the authentic protease. This is the first example of the direct secretion of a Gram- extracellular enzyme in B. subtilis via its own signal peptide. The fact that this gene can be expressed and its product secreted in both Escherichia coli and B. subtilis provides a unique opportunity to study and compare the similarities and differences in protein secretion between Gram- and Gram+ organisms.
Wang
L F
LF
CSIRO Australian Animal Health Laboratory, Geelong, Victoria, Australia.
Kortt
A A
AA
Stewart
D J
DJ
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology*; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D001439:Bacteroides / Q000201:enzymology*; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D005798:Genes, Bacterial; D006090:Gram-Negative Bacteria / Q000201:enzymology* / Q000235:genetics; D006094:Gram-Positive Bacteria / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D010957:Plasmids; D021382:Protein Sorting Signals / Q000502:physiology*; D011993:Recombinant Fusion Proteins; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D013045:Species Specificity; D013381:Subtilisins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D012697:Serine Endopeptidases; D013381:Subtilisins; C078345:extracellular basic protease, Dichelobacter nodosus