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.

Functional characterization of a gene locus from an uncultured gut Bacteroides conferring xylo‐oligosaccharides utilization to Escherichia coli

“…As showed here, the direct study of metagenomic clones for characterization of new functions and/or new protein families could be a good strategy not only to save time, but also to study some complex mechanisms that require the synergistic action of different proteins. Previously, some multigenic systems issued of metagenomic libraries have been used to optimize E. coli abilities to produce ethanol or antifungal activity (Chung et al ., ; Loaces et al ., ), but no biochemical characterization of PULs heterologously expressed in E. coli has been published so far. Here, we demonstrated that E. coli is an interesting recombinant host for characterizing the individual components of PUL systems from Bacteroides strains.…”

A common vaginal microbiota composition among breeds of Bos taurus indicus (Gyr and Nellore)

“…The study of the microbiota has provided important insights into health and pathology of animals of economic importance [1]. To date, most of the research has focused on the ruminal microbiota due its biotechnological interest [2] and in order to understand and reduce the pressing environmental problems associated with greenhouse gas emissions from ruminant livestock production [3,4]. The economic importance of cattle, as well as the need for a better understanding of resident microbial communities, has led to the development of many studies to elucidate the relationship of the indigenous microbiota with the physiology of these animals.…”

Indole modifies the central carbon flux in the anaerobic metabolism of Escherichia coli: application to the production of hydrogen and other metabolites

“…E. coli is currently used as a model for production studies of hydrogen, ethanol and other metabolites, since its anaerobic metabolism is well documented and genetic manipulations are possible. Both genetic modifications and modified fermentation strategies have been implemented to improve the production processes [7,13,14]. Since the production of metabolites normally correlates with cell growth, an important fraction of the substrate is inevitably channeled toward unwanted biomass rather than the desired product.…”