Properties of polypeptides secreted into the growth medium by the Ftp library clones and purified His-recombinant polypeptides. A. Upper panel shows the binding of cell-free growth media from the library clones to ECM proteins and the control protein fetuin immobilized in polystyrene microtitre wells as analyzed by ELISA. Lower panel shows Western blot analysis with monoclonal anti-FLAG antibodies of bacterial cells (C) and TCA-precipitated cell-free growth media (S) of the corresponding clones. Vector indicates growth medium from MKS12 (pSRP18/0), D1-D3 denotes polypeptides secreted by MKS12 (pSRP18/0D1-D3), and the names indicate individual library clones. The cell samples correspond to 50 μl and the supernatants to 500 μl bacterial culture, except in the case of clones ΔPBP, ΔUsp, ΔIspD, ΔEbh where supernatant samples corresponding only to 100 μl of culture were loaded due to the high expression level of the polypeptide. B. Upper panel presents the binding of His-tagged recombinant polypeptides to ECM proteins immobilized in polystyrene microtiter wells as analyzed by ELISA and the lower panel shows SDS-PAGE analysis of affinity-purified recombinant polypeptides. The names following His-indicate polypeptides encoded by gene fragments subcloned from corresponding individual library clones. The values are averages of 2 to 3 parallels from 2 to 4 individual experiments, showing the standard deviation as error bars. CI, type I collagen; CIV, type IV collagen; Fn, fibronectin; Fg, fibrinogen; Fet, control protein fetuin. Molecular masses in kDa are indicated to the left.
Kylväjä et al. BMC Microbiology 2011 11:117 doi:10.1186/1471-2180-11-117