Figure 5

Covalent binding is caused by interaction of SpyCatcher with SpyTag. The table at the top indicates the approximate expected sizes of conjugated and unconjugated proteins. Bottom: duplicate anti-HBcAg (left) and anti-GFP (right) western blots of total soluble protein obtained from leaves co-expressing the proteins indicated. Higher molecular weight multimers of tandem core proteins are visible in the anti-HBcAg blot for all constructs, but these are only visible in the anti-GFP blot when the SpyCatcher/SpyTag interaction can take place (first lane), indicating covalent binding of GFP to HBcAg. Monomers of GFP-ST and GFP-SC are visible in the anti-GFP blot (arrows 5 and 6, respectively) along with bands of an unknown nature (marked “?”) which cannot be GFP bound to tHBcAg since these bands are not present on the anti-HBcAg blot. Blots cropped for clarity, for uncropped blots see Supplementary Fig. S4.