System Using Tandem Repeats of the cA Peptidoglycan-Binding Domain from Lactococcus lactis for Display of both N- and C-Terminal Fusions on Cell Surfaces of Lactic Acid Bacteria

Department of Molecular Science and Material Engineering, Graduate School of Science and Technology, Kobe University, 1-1 Rokkodaicho, Nada-ku, Kobe 657-8501, Japan.
Applied and Environmental Microbiology (Impact Factor: 3.67). 03/2008; 74(4):1117-23. DOI: 10.1128/AEM.02012-07
Source: PubMed


Here, we established a system for displaying heterologous protein to the C terminus of the peptidoglycan-binding domain (cA domain) of AcmA (a major autolysin from Lactococcus lactis). Western blot and flow cytometric analyses revealed that the fusion proteins (cA-AmyA) of the cA domain and alpha-amylase from Streptococcus bovis 148 (AmyA) are efficiently expressed and successfully displayed on the surfaces of L. lactis cells. AmyA was also displayed on the cell surface while retaining its activity. Moreover, with an increase in the number of cA domains, the quantity of cA-AmyA fusion proteins displayed on the cell surface increased. When three repeats of the cA domain were used as an anchor protein, 82% of alpha-amylase activity was detected on the cells. The raw starch-degrading activity of AmyA was significantly higher when AmyA was fused to the C terminus of the cA domain than when it was fused to the N terminus. In addition, cA-AmyA fusion proteins were successfully displayed on the cell surfaces of Lactobacillus plantarum and Lactobacillus casei.

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    • "Surface-anchored complexes were thus displayed with efficiencies that could approach 10 4 complexes/cell (Wieczorek and Martin, 2010) (Table 1). A non-covalent surface display system for LAB has also been developed by fusing a target heterologous protein, i.e., the α-amylase, with the C-terminal cA peptidoglycan-binding domain, which shows high homology with LysM repeats of the major autolysin AcmA from Lc. lactis (Okano et al., 2008). "
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