Monday, August 11, 2008 - 10:50 AM
S18

In-vivo assembly of cell-surface associated cellulosome-inspired supramolecular scaffolds in Lactococcus lactis

Andrew S. Wieczorek and Vincent J.J. Martin. Biology, Concordia University, 7141 Sherbrooke St. West, Montreal, QC H4B1R6, Canada

The production of microbial protein scaffolds for the ordered display of synergistic enzymes provides a template for the assembly of industrially relevant and novel molecules by means of a veritable microbial “assembly line”. Clostridium thermocellum is an anaerobic bacterium capable of hydrolyzing cellulosic substrates by means of a highly active and thermally stable, cell-associated multi-enzyme complex termed the cellulosome. Fragments of the major component CipA, an enzyme-binding scaffold, were successfully anchored onto a surrogate host, Lactococcus lactis. Chimeric scaffolds were fused to an N-terminal signal peptide for secretion as well as a C-terminal anchor domain, and were expressed under the control an inducible promoter. A total of 5 chimeric scaffolds were successfully displayed on the cell-surface, and 5 reporters displaying either fluorescent or enzymatic activity were engineered to associate with the scaffolds by means of a C-terminal “dockerin domain”, forming a strong non-covalent interaction with one of the scaffold’s “cohesin domains”. This “plug-and-play” option thus allows the assembly of enzymes and reporters onto the cell-bound microbial scaffold in an ordered fashion. Such cell-associated enzymatic complexes consisting of synergistic enzymes of choice have endless potential for use in the assembly of industrially relevant molecules, the decomposition of larger polymers, and for studying the synergistic effects of enzymes when assembled within variable proximity to one another.

See more of Student Oral Presentations
See more of Invited Oral Papers

See more of The Annual Meeting and Exhibition 2008 (August 10 - 14, 2008)