title: Improved ethanol production from biomass by a rumen metagenomic DNA fragment expressed in Escherichia coli MS04 during fermentation.
abstract: With the aim of improving current ethanologenic Escherichia coli strains, we screened a metagenomic library from bovine ruminal fluid for cellulolytic enzymes. We isolated one fosmid, termed Csd4, which was able to confer to E. coli the ability to grow on complex cellulosic material as the sole carbon source such as avicel, carboxymethyl cellulose, filter paper, pretreated sugarcane bagasse, and xylan. Glucanolytic activity obtained from E. coli transformed with Csd4 was maximal at 24 h of incubation and was inhibited when glucose or xylose were present in the media. The 34,406-bp DNA fragment of Csd4 was completely sequenced, and a putative endoglucanase, a xylosidase/arabinosidase, and a laccase gene were identified. Comparison analysis revealed that Csd4 derived from an organism closely related to Prevotella ruminicola, but no homologies were found with any of the genomes already sequenced. Csd4 was introduced into the ethanologenic E. coli MS04 strain and ethanol production from CMC, avicel, sugarcane bagasse, or filter paper was observed. Exogenously expressed β-glucosidase had a positie effect on cell growth in agreement with the fact that no putative β-glucosidase was found in Csd4. Ethanol production from sugarcane bagasse was improved threefold by Csd4 after saccharification by commercial Trichoderma reesei cellulases underlining the ability of Csd4 to act as a saccharification enhancer to reduce the enzymatic load and time required for cellulose deconstruction.
authors.affiliation: Departamento de Bioquímica y Genómica Microbianas, Instituto de Investigaciones Biológicas Clemente Estable, Av. Italia 3318, Montevideo, 11600, Uruguay.
authors.forename: Inés
authors.initials: I
authors.lastname: Loaces
authors.affiliation: Departamento de Bioquímica y Genómica Microbianas, Instituto de Investigaciones Biológicas Clemente Estable, Av. Italia 3318, Montevideo, 11600, Uruguay.
authors.forename: Vanesa
authors.initials: V
authors.lastname: Amarelle
authors.affiliation: Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, A.P: 510-3., Cuernavaca, Morelos, 62270, México.|Department of Molecular Microbiology and Biotechnology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Iván
authors.initials: I
authors.lastname: Muñoz-Gutierrez
authors.affiliation: Departamento de Bioquímica y Genómica Microbianas, Instituto de Investigaciones Biológicas Clemente Estable, Av. Italia 3318, Montevideo, 11600, Uruguay.
authors.forename: Elena
authors.initials: E
authors.lastname: Fabiano
authors.affiliation: Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, A.P: 510-3., Cuernavaca, Morelos, 62270, México.
authors.forename: Alfredo
authors.initials: A
authors.lastname: Martinez
authors.affiliation: Departamento de Bioquímica y Genómica Microbianas, Instituto de Investigaciones Biológicas Clemente Estable, Av. Italia 3318, Montevideo, 11600, Uruguay. fnoya@iibce.edu.uy.
authors.forename: Francisco
authors.initials: F
authors.lastname: Noya
mesh_terms: D000818:Animals; D018533:Biomass; D001711:Biotransformation; D002417:Cattle; D002480:Cellulase / Q000235:genetics; D002482:Cellulose / Q000378:metabolism; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D000431:Ethanol / Q000378:metabolism*; D005285:Fermentation; D015870:Gene Expression; D042845:Laccase / Q000235:genetics; D060847:Metabolic Engineering; D054892:Metagenome; D045243:Prevotella ruminicola / Q000235:genetics; D012417:Rumen / Q000382:microbiology*; D031786:Saccharum / Q000737:chemistry; D017422:Sequence Analysis, DNA; D014995:Xylosidases / Q000235:genetics
chemical_list: D000431:Ethanol; D002482:Cellulose; D004247:DNA; D042845:Laccase; D014995:Xylosidases; D002480:Cellulase
keywords: Bioethanol; Biomass hydrolysis; CBP; Cellulolytic enzymes; Rumen metagenomics