Expression of the Staphylococcus hyicus lipase in Lactococcus lactis.
The extracellular Staphylococcus hyicus lipase was expressed under the control of different promoters in Lactococcus lactis and Bacillus subtilis. Its expression at high and moderate levels is toxic for the former and the latter hosts, respectively. In L. lactis, the lipase was expressed at a high level, up to 30% of the total cellular proteins, under the control of the inducible promoter PnisA. About 80% of the lipase remained associated with the cells. Close to half of this amount remained associated with the inner side of the cytoplasmic membrane as unprocessed pre-pro-lipase. The other half was trapped by the cell wall and partially degraded at the N-terminal end. This result suggests that extracellular proteases degrade the lipase. Surprisingly, the kinetics and the pattern of lipase degradation were different in the two L. lactis subspecies, L. lactis subsp. cremoris and L. lactis subsp. lactis. The extracellular proteolytic systems that degrade lipase are thus different in these closely related subspecies. The incorrect export of the lipase is not due to an inappropriate leader peptide but may be due to an inefficiency of several steps of lipase secretion. We propose that (i) the S. hyicus lipase may require a special accessory system to be correctly exported or (ii) the kinetics of lipase synthesis may be a critical factor for proper folding.
Drouault
S
S
Unité de Génétique Microbienne, Institut National de la Recherche Agronomique, 78352 Jouy en Josas Cedex, France.
Corthier
G
G
Ehrlich
S D
SD
Renault
P
P
D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D015153:Blotting, Western; D002458:Cell Fractionation; D013294:Lactococcus lactis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D008049:Lipase / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D013210:Staphylococcus / Q000201:enzymology / Q000235:genetics*
D011994:Recombinant Proteins; D008049:Lipase