title: Construction of cellulose-utilizing Escherichia coli based on a secretable cellulase.
abstract: BACKGROUND
  The microbial conversion of plant biomass into value added products is an attractive option to address the impacts of petroleum dependency. The Gram-negative bacterium Escherichia coli is commonly used as host for the industrial production of various chemical products with a variety of sugars as carbon sources. However, this strain neither produces endogenous cellulose degradation enzymes nor secrets heterologous cellulases for its poor secretory capacity. Thus, a cellulolytic E. coli strain capable of growth on plant biomass would be the first step towards producing chemicals and fuels. We previously identified the catalytic domain of a cellulase (Cel-CD) and its N-terminal sequence (N20) that can serve as carriers for the efficient extracellular production of target enzymes. This finding suggested that cellulose-utilizing E. coli can be engineered with minimal heterologous enzymes.
  
  
  RESULTS
  In this study, a β-glucosidase (Tfu0937) was fused to Cel-CD and its N-terminal sequence respectively to obtain E. coli strains that were able to hydrolyze the cellulose. Recombinant strains were confirmed to use the amorphous cellulose as well as cellobiose as the sole carbon source for growth. Furthermore, both strains were engineered with poly (3-hydroxybutyrate) (PHB) synthesis pathway to demonstrate the production of biodegradable polyesters directly from cellulose materials without exogenously added cellulases. The yield of PHB reached 2.57-8.23 wt% content of cell dry weight directly from amorphous cellulose/cellobiose. Moreover, we found the Cel-CD and N20 secretion system can also be used for the extracellular production of other hydrolytic enzymes.
  
  
  CONCLUSIONS
  This study suggested that a cellulose-utilizing E. coli was created based on a heterologous cellulase secretion system and can be used to produce biofuels and biochemicals directly from cellulose. This system also offers a platform for conversion of other abundant renewable biomass to biofuels and biorefinery products.
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, People's Republic of China. ga_o_df@126.com.
authors.forename: Dongfang
authors.initials: D
authors.lastname: Gao
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, People's Republic of China. ytluanyaqi@126.com.
authors.forename: Yaqi
authors.initials: Y
authors.lastname: Luan
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, People's Republic of China. qiqi20011983@gmail.com.
authors.forename: Qian
authors.initials: Q
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, People's Republic of China. liangquanfeng@sdu.edu.cn.
authors.forename: Quanfeng
authors.initials: Q
authors.lastname: Liang
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, People's Republic of China. qiqingsheng@sdu.edu.cn.
authors.forename: Qingsheng
authors.initials: Q
authors.lastname: Qi
mesh_terms: D039903:Actinobacteria / Q000201:enzymology; D018533:Biomass; D002480:Cellulase / Q000235:genetics / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D005947:Glucose / Q000378:metabolism; D006885:Hydroxybutyrates / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011091:Polyesters / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D000094122:Polyhydroxybutyrates
chemical_list: D002480:Cellulase; D002482:Cellulose; D005947:Glucose; D006885:Hydroxybutyrates; D011091:Polyesters; D011993:Recombinant Fusion Proteins; C003182:poly-beta-hydroxybutyrate; D000094122:Polyhydroxybutyrates