Figure 4
Multimerization triggered by the HC. (A) Western blotting of the secreted recombinant VLRBs. The secretion of the randomly selected rVLRBs (rVLRB#1, rVLRB#2) was visualized by Western blotting with 11G5 after running on nave-PAGE (8%) or SDS-PAGE (12%). (B) Western blotting of RFP-stalk or RFP-stalk-HC. Each supernatant (5 and 0.5 μg for RFP-stalk; 5 and 100 μg for RFP-stalk-HC) was separated by 8% native-PAGE under non-reducing condition, followed by Western blot analysis with 11G5. The original full-length blot is presented in Supplementary Fig. S2. (C) Minimum requirement of the HC8 for multimerization of RFP-stalk. Supernatants from 293-F cells transfected with the plasmids expressing RFP-stalk with or without various versions of the HC (HC5, HC6, HC7, HC8, HC9, HC10, HC15, HC20, HC25, or HC; see Supplementary Table S1) were run on 8% native-PAGE, and subjected to Western blot analysis with 11G5. (D) Reduced expression levels depending on the sizes of the conjugated HC. Each supernatant was separated by 12% SDS-PAGE under non-reducing conditions followed by Western blotting with 11G5.
