In vivo binding of UidAdock1 on live intact L. lactis cells displaying CipAfrags. CipAfrags were expressed and anchored as fusions with the NucA reporter enzyme (A), or lacking the NucA reporter (B). Quantification of UidAdock1 molecules bound to L. lactis cells corresponds to equivalent amounts of functional cohesin assuming a 1:1 ratio of dockerin-cohesin association. Dark grey bars represent scaffolds containing the C-terminal M6 cell wall anchor motif (cwa), and light grey bars represent their anchor-deficient derivatives. White bars correspond to indicated controls; "200 μg/mL UidA-dock1" represents binding assay carried out with excess enzyme and L. lactis pAW328 (NucA-CBM3a-coh3-cwa) to ensure saturation of cohesins. "100 μg/mL UidA" represents binding assay carried out in the presence of UidA and L. lactis pAW328 (NucA-CBM3a-cwa). Binding assay carried out with UidA and all other constructs resulted in no association with scaffold-expressing strains (data not shown).
Wieczorek and Martin Microbial Cell Factories 2010 9:69 doi:10.1186/1475-2859-9-69