title: Optimization of protease secretion in Bacillus subtilis and Bacillus licheniformis by screening of homologous and heterologous signal peptides.
abstract: Bacillus subtilis and Bacillus licheniformis are widely used for the large-scale industrial production of proteins. These strains can efficiently secrete proteins into the culture medium using the general secretion (Sec) pathway. A characteristic feature of all secreted proteins is their N-terminal signal peptides, which are recognized by the secretion machinery. Here, we have studied the production of an industrially important secreted protease, namely, subtilisin BPN' from Bacillus amyloliquefaciens. One hundred seventy-three signal peptides originating from B. subtilis and 220 signal peptides from the B. licheniformis type strain were fused to this secretion target and expressed in B. subtilis, and the resulting library was analyzed by high-throughput screening for extracellular proteolytic activity. We have identified a number of signal peptides originating from both organisms which produced significantly increased yield of the secreted protease. Interestingly, we observed that levels of extracellular protease were improved not only in B. subtilis, which was used as the screening host, but also in two different B. licheniformis strains. To date, it is impossible to predict which signal peptide will result in better secretion and thus an improved yield of a given extracellular target protein. Our data show that screening a library consisting of homologous and heterologous signal peptides fused to a target protein can identify more-effective signal peptides, resulting in improved protein export not only in the original screening host but also in different production strains.
authors: Institute of Molecular Enzyme Technology, Heinrich-Heine-Universität Düsseldorf, Research Center Jülich, Jülich, Germany.
authors: Christian
authors: C
authors: Degering
authors: Thorsten
authors: T
authors: Eggert
authors: Michael
authors: M
authors: Puls
authors: Johannes
authors: J
authors: Bongaerts
authors: Stefan
authors: S
authors: Evers
authors: Karl-Heinz
authors: KH
authors: Maurer
authors: Karl-Erich
authors: KE
authors: Jaeger
mesh_terms: D001407:Bacillus / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001709:Biotechnology / Q000379:methods; D007218:Industrial Microbiology; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013381:Subtilisins / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D013381:Subtilisins