Abstract
Optimising the production of heterologous exported proteins in Lactococcus lactis by inactivation of HtrA, the unique housekeeping surface protease. The use of bacteria as cell fac- tories to produce heterologous exported proteins is often limited by extra-cellular proteolysis. We con- structed a Lactococcus lactis mutant strain in which recombinant or heterologous exported proteins are stable. A previously unknown membrane protease belonging to the HtrA/DegP family (HtrA LI ) was first identified in L. lactis subsp. lactis strain IL1403. Inactivation of the chromosomal gene revealed that HtrA LI acts as a surface housekeeping protease by elimination of abnormal and/or mis- folded proteins, and that HtrA LI is also responsible for the maturation of natural exported proteins, such as the L. lactis bacteriolysin, AcmA. From an applied point of view, the most important result of our study is that in the absence of HtrA LI , the extra-cellular proteolysis of all tested proteins, in particu- lar an heterologous one, is completely abolished, and the yield of intact protein is significantly increased. These results suggest that HtrA LI is the sole extra-cellular housekeeping protease in L. lactis, in agreement with the analysis of the complete IL1403 genome sequence. In the future, the mutant htrA strain should constitute an efficient tool to improve yields of heterologous exported proteins in L. lactis. Lactococcus lactis HtrA / extra-cellular proteolysis / extra-cellular protein stability / production of heterologous proteins
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