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Efficient production of secreted staphylococcal antigens in a non-lysing and proteolytically reduced Lactococcus lactis strain

Neef, J., Koedijk, D. G. A. M., Bosma, T., van Dijl, J. M. & Buist, G., Dec-2014, In : Applied Microbiology and Biotechnology. 98, 24, p. 10131-10141 11 p.

Research output: Contribution to journalArticleAcademicpeer-review

Cell surface-exposed and secreted proteins are attractive targets for vaccination against pathogenic gram-positive bacteria. To obtain sufficient amounts of such antigens, efficient protein production platforms are needed. In this study, a pipeline for the production and purification of surface-exposed and secreted antigens of the gram-positive bacterial pathogen Staphylococcus aureus is presented. Cytoplasmic or extracellular production of S. aureus antigens was achieved using the Lactococcus lactis strain PA1001, which lacks the major extracellular protease HtrA and the autolysin AcmA to minimize proteolysis and cell lysis, respectively. For most tested S. aureus antigens, secretory production directed by the signal peptide of the major secreted protein Usp45 of L. lactis resulted in higher yields than intracellular production without a signal peptide. Additionally, secretory production of His-tagged antigens allowed their facile one-step purification from the growth medium by metal affinity chromatography. For three of the purified antigens, biological activity was confirmed through enzyme activity assays. We, furthermore, show that the present pipeline can be used to produce staphylococcal antigens with an N-terminal AVI-tag for site-specific labeling with biotin or a C-terminal cell wall-binding domain for cell surface display. We conclude that our L. lactis-based pipeline allows the efficient production of S. aureus antigens and their subsequent purification in one step.

Original languageEnglish
Pages (from-to)10131-10141
Number of pages11
JournalApplied Microbiology and Biotechnology
Volume98
Issue number24
Publication statusPublished - Dec-2014

    Keywords

  • Lactococcus lactis, Staphylococcus aureus, Antigens, Secretion, One-step purification, CONTROLLED GENE-EXPRESSION, HETEROLOGOUS PROTEINS, BACILLUS-SUBTILIS, AUREUS, PROTEOMICS, SYSTEM, NISIN, VACCINE, CONSTRUCTION, INFECTIONS

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