title: Engineering a homobutanol fermentation pathway in Escherichia coli EG03.
abstract: A homobutanol fermentation pathway was engineered in a derivative of Escherichia coli B (glucose [glycolysis] => 2 pyruvate + 2 NADH; pyruvate [pyruvate dehydrogenase] => acetyl-CoA + NADH; 2 acetyl-CoA [butanol pathway enzymes] + 4 NADH => butanol; summary stoichiometry: glucose => butanol). Initially, the native fermentation pathways were eliminated from E. coli B by deleting the genes encoding for lactate dehydrogenase (ldhA), acetate kinase (ackA), fumarate reductase (frdABCD), pyruvate formate lyase (pflB), and alcohol dehydrogenase (adhE), and the pyruvate dehydrogenase complex (aceEF-lpd) was anaerobically expressed through promoter replacement. The resulting strain, E. coli EG03 (ΔfrdABCD ΔldhA ΔackA ΔpflB Δ adhE ΔpdhR ::pflBp6-aceEF-lpd ΔmgsA), could generate 4 NADH for every glucose oxidized to two acetyl-CoA through glycolysis and the pyruvate dehydrogenase complex. However, EG03 lost its ability for anaerobic growth due to the lack of NADH oxidation pathways. When the butanol pathway genes that encode for acetyl-CoA acetyltransferase (thiL), 3-hydroxybutyryl-CoA dehydrogenase (hbd), crotonase (crt), butyryl-CoA dehydrogenase (bcd, etfA, etfB), and butyraldehyde dehydrogenase (adheII) were cloned from Clostridium acetobutylicum ATCC 824, and expressed in E. coli EG03, a balanced NADH oxidation pathway was established for homobutanol fermentation (glucose => 4 NADH + 2 acetyl-CoA => butanol). This strain was able to convert glucose to butanol (1,254 mg l(-1)) under anaerobic condition.
authors: Key Laboratory of Fermentation Engineering (Ministry of Education), College of Bioengineering, Hubei University of Technology, Wuhan, 430068, People's Republic of China.
authors: Erin
authors: E
authors: Garza
authors: Jinfang
authors: J
authors: Zhao
authors: Yongze
authors: Y
authors: Wang
authors: Jinhua
authors: J
authors: Wang
authors: Andrew
authors: A
authors: Iverson
authors: Ryan
authors: R
authors: Manow
authors: Chris
authors: C
authors: Finan
authors: Shengde
authors: S
authors: Zhou
mesh_terms: D020001:1-Butanol / Q000378:metabolism*; D015094:3-Hydroxyacyl CoA Dehydrogenases / Q000235:genetics / Q000378:metabolism; D000101:Acetyl-CoA C-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D000426:Alcohol Dehydrogenase / Q000235:genetics / Q000378:metabolism; D056804:Biofuels; D019149:Bioreactors; D000440:Butanols / Q000378:metabolism*; D044926:Butyryl-CoA Dehydrogenase / Q000235:genetics / Q000378:metabolism; D046969:Clostridium acetobutylicum / Q000201:enzymology / Q000235:genetics; D004746:Enoyl-CoA Hydratase / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D005285:Fermentation; D005947:Glucose / Q000378:metabolism; D006019:Glycolysis; D060847:Metabolic Engineering; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D011768:Pyruvate Dehydrogenase Complex / Q000235:genetics / Q000378:metabolism
chemical_list: D056804:Biofuels; D000440:Butanols; D011768:Pyruvate Dehydrogenase Complex; D009243:NAD; D020001:1-Butanol; D015094:3-Hydroxyacyl CoA Dehydrogenases; D000426:Alcohol Dehydrogenase; C021388:3-hydroxybutyryl-CoA dehydrogenase; D044926:Butyryl-CoA Dehydrogenase; D000101:Acetyl-CoA C-Acetyltransferase; D004746:Enoyl-CoA Hydratase; D005947:Glucose