title: Enhanced cellulose degradation by nano-complexed enzymes: Synergism between a scaffold-linked exoglucanase and a free endoglucanase.
abstract: Protein molecular scaffolds are attracting interest as natural candidates for the presentation of enzymes and acceleration of catalytic reactions. We have previously reported evidence that the stable protein 1 (SP1) from Populustremula can be employed as a molecular scaffold for the presentation of either catalytic or structural binding (cellulosomal cohesin) modules. In the present work, we have displayed a potent exoglucanase (Cel6B) from the aerobic cellulolytic bacterium, Thermobifida fusca, on a cohesin-bearing SP1 scaffold. For this purpose, a chimaeric form of the enzyme, fused to a cellulosomal dockerin module, was prepared. Full incorporation of 12 dockerin-bearing exoglucanase molecules onto the cohesin-bearing scaffold was achieved. Cellulase activity was tested on two cellulosic substrates with different levels of crystallinity, and the activity of the scaffold-linked exoglucanase was significantly reduced, compared to the free dockerin-containing enzyme. However, addition of relatively low concentrations of a free wild-type endoglucanase (T. fusca Cel5A) that bears a cellulose-binding module, in combination with the complexed exoglucanase resulted in a marked rise in activity on both cellulosic substrates. The endoglucanase cleaves internal sites of the cellulose chains, and the new chain ends of the substrate were now readily accessible to the scaffold-borne exoglucanase, thereby resulting in highly effective, synergistic degradation of cellulosic substrates.
authors.affiliation: Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Sarah
authors.initials: S
authors.lastname: Moraïs
authors.forename: Arnon
authors.initials: A
authors.lastname: Heyman
authors.forename: Yoav
authors.initials: Y
authors.lastname: Barak
authors.forename: Jonathan
authors.initials: J
authors.lastname: Caspi
authors.forename: David B
authors.initials: DB
authors.lastname: Wilson
authors.forename: Raphael
authors.initials: R
authors.lastname: Lamed
authors.forename: Oded
authors.initials: O
authors.lastname: Shoseyov
authors.forename: Edward A
authors.initials: EA
authors.lastname: Bayer
mesh_terms: D001709:Biotechnology / Q000379:methods*; D018797:Cell Cycle Proteins / Q000378:metabolism; D002480:Cellulase / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism*; D002850:Chromatography, Gel; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D003013:Clostridium / Q000378:metabolism; D049329:Nanostructures / Q000737:chemistry* / Q000648:ultrastructure; D010940:Plant Proteins / Q000648:ultrastructure; D032107:Populus / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D000097722:Cohesins
chemical_list: D018797:Cell Cycle Proteins; D002480:Cellulase; D002482:Cellulose; D002868:Chromosomal Proteins, Non-Histone; D010940:Plant Proteins; D011994:Recombinant Proteins; D000097722:Cohesins