title: Significance of relative position of cellulases in designer cellulosomes for optimized cellulolysis.
abstract: Degradation of cellulose is of major interest in the quest for alternative sources of renewable energy, for its positive effects on environment and ecology, and for use in advanced biotechnological applications. Due to its microcrystalline organization, celluose is extremely difficult to degrade, although numerous microbes have evolved that produce the appropriate enzymes. The most efficient known natural cellulolytic system is produced by anaerobic bacteria, such as C. thermocellum, that possess a multi-enzymatic complex termed the cellulosome. Our laboratory has devised and developed the designer cellulosome concept, which consists of chimaeric scaffoldins for controlled incorporation of recombinant polysaccharide-degrading enzymes. Recently, we reported the creation of a combinatorial library of four cellulosomal modules comprising a basic chimaeric scaffoldin, i.e., a CBM and 3 divergent cohesin modules. Here, we employed selected members of this library to determine whether the position of defined cellulolytic enzymes is important for optimized degradation of a microcrystalline cellulosic substrate. For this purpose, 10 chimaeric scaffoldins were used for incorporation of three recombinant Thermobifida fusca enzymes: the processive endoglucanase Cel9A, endoglucanase Cel5A and exoglucanase Cel48A. In addition, we examined whether the characteristic properties of the T. fusca enzymes as designer cellulosome components are unique to this bacterium by replacing them with parallel enzymes from Clostridium thermocellum. The results support the contention that for a given set of cellulosomal enzymes, their relative position within a scaffoldin can be critical for optimal degradation of microcrystaline cellulosic substrates.
authors.affiliation: Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Johanna
authors.initials: J
authors.lastname: Stern
authors.affiliation: Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Amaranta
authors.initials: A
authors.lastname: Kahn
authors.affiliation: Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Yael
authors.initials: Y
authors.lastname: Vazana
authors.affiliation: Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Melina
authors.initials: M
authors.lastname: Shamshoum
authors.affiliation: Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Sarah
authors.initials: S
authors.lastname: Moraïs
authors.affiliation: Department of Molecular Microbiology and Biotechnology, Tel Aviv University, Ramat Aviv 69978, Israel.
authors.forename: Raphael
authors.initials: R
authors.lastname: Lamed
authors.affiliation: Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Edward A
authors.initials: EA
authors.lastname: Bayer
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D002480:Cellulase / Q000737:chemistry / Q000235:genetics; D048013:Acetivibrio thermocellus / Q000201:enzymology / Q000235:genetics; D015723:Gene Library; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D002480:Cellulase