The type II cohesin domain and type II dockerin of bacterial cellulosome were cloned from Clostridium thermocellum and expressed with the fusion of tobacco mosaic virus coat protein (TMVcp) and enhanced green fluorescent protein (EGFP), respectively, in Escherichia coli. The TMVcp-cohesin fusion protein was assembled to the stable and rod-shaped nanostructure (TMVcp-Coh rod) under a particular buffer condition, where many active cohesin proteins are biologically and densely displayed around the 3-dimensional surface of TMVcp-Coh rod. Using EGFP-dockerin as a fluorescent reporter, we confirmed that the Ca2+-dependent binding and dissociation between native cohesin and dockerin were reproduced with the two recombinant fusion proteins, TMVcp-cohesin and EGFP-dockerin. The multi-cyclic binding-dissociation operation of TMVcp-Coh rod and EGFP-dockerin was successfully performed with maintaining the reversible cohesin-dockerin interaction in every cycle. EGFP that was fused to dockerin as a proof-of-concept here can be switched to other functional proteins/peptides that need to be used in multi-cyclic operation.
Bibliographical noteFunding Information:
This study was supported by the 2015 NLRL (National Leading Research Lab.) Project (grant no. 2015R1A2A1A05001861 ) (the main project that supported this work), the Basic Science Research Program (ERC program, grant no. 2010-0027955 ), and the 2013 NLRL Project (grant no. 2013R1A2A1A01015644 ) of the National Research Foundation of Korea (NRF).
© 2016 Elsevier B.V.
- Multi-cyclic operation
- Reversible interaction
- Tobacco mosaic virus coat protein
ASJC Scopus subject areas
- Applied Microbiology and Biotechnology