Cell surface display of a β-glucosidase employing the type V secretion system on ethanologenic Escherichia coli for the fermentation of cellobiose to ethanol

We used the autodisplay system AIDA-I, which belongs to the type V secretion system (TVSS), to display the beta-glucosidase BglC from Thermobifida fusca on the outer membrane of the ethanologenic Escherichia coli strain MS04 (MG1655 a dagger pflB, a dagger adhE, a dagger frdA, a dagger xylFGH, a dagger ldhA, PpflB::pdc (Zm) -adhB (Zm) ). MS04 that was transformed with the plasmid pAIDABglCRHis showed cellobiase activity (171 U/g(CDW)) and fermented 40 g/l cellobiose in mineral medium in 60 h with an ethanol yield of 81 % of the theoretical maximum. Whole-cell protease treatment, SDS-PAGE, and Western-blot analysis demonstrated that BglC was attached to the external surface of the outer membrane of MS04. When attached to the cells, BglC showed 93.3 % relative activity in the presence of 40 g/l ethanol and retained 100 % of its activity following 2 days of incubation at 37 A degrees C with the same ethanol concentration. This study shows the potential of the TVSS (AIDA-I) and BglC as tools for the production of lignocellulosic bio-commodities.