Fig. 3

Optimization of the CatIB strategy by variation of coiled-coil domains using 3HAMP instead of TDoT fusions. CatIB formation was evaluated by a SDS-PAGE analysis and b CatIB formation efficiency for 3HAMP-L-LbADH- and 3HAMP-L-PpBFD (green bars) compared to TDoT-L-LbADH and TDoT-L-PpBFD (blue bars). Panels c) and d) contain the equivalent data for 3HAMP-L-YFP, 3HAMP-L-mCherry, 3HAMP-L-PfBAL and 3HAMP-L-RADH. After cell disruption, the crude cell extract (CCE) was separated by centrifugation into the soluble protein containing supernatant (SN) and the insoluble IB containing pellet (P) fraction. Sample preparation for SDS-PAGE analysis and the determination of the CatIB/FIB formation efficiency was carried out as described in Fig. 1. a, c SDS-PAGE analysis of the respective protein fractions: CCE, SN, and P. The molecular mass of the target fusion proteins is indicated by arrows (3HAMP-L-LbADH: 47.1 kDa; 3HAMP-L-PpBFD: 77.0 kDa, 3HAMP-L-YFP: 47.4 kDa, 3HAMP-L-mCherry: 47.1 kDa, 3HAMP-L-RADH: 47.1 kDa and 3HAMP-L-PfBAL: 79.3 kDa). b, d CatIB/FIB formation efficiency determined as described in Fig. 1. The complete datasets illustrating the distribution of activity in the CCE, SN and P fractions can be found in Additional file 1: Figure S7. Initial rate activities were measured as described in Fig. 1