Production of C4 and C5 branched-chain alcohols by engineered Escherichia. coli.
Higher alcohols, longer chain alcohols, contain more than 3 carbon atoms, showed close energy advantages as gasoline, and were considered as the next generation substitution for chemical fuels. Higher alcohol biosynthesis by native microorganisms mainly needs gene expression of heterologous keto acid decarboxylase and alcohol dehydrogenases. In the present study, branched-chain α-keto acid decarboxylase gene from Lactococcus lactis subsp. lactis CICC 6246 (Kivd) and alcohol dehydrogenases gene from Zymomonas mobilis CICC 41465 (AdhB) were transformed into Escherichia coli for higher alcohol production. SDS-PAGE results showed these two proteins were expressed in the recombinant strains. The resulting strain was incubated in LB medium at 37 °C in Erlenmeyer flasks and much more 3-methyl-1-butanol (104 mg/L) than isobutanol (24 mg/L) was produced. However, in 5 g/L glucose-containing medium, the production of two alcohols was similar, 156 and 161 mg/L for C4 (isobutanol) and C5 (3-methyl-1-butanol) alcohol, respectively. Effects of fermentation factors including temperature, glucose content, and α-keto acid on alcohol production were also investigated. The increase of glucose content and the adding of α-keto acids facilitated the production of C4 and C5 alcohols. The enzyme activities of pure Kivd on α-ketoisovalerate and α-ketoisocaproate were 26.77 and 21.24 μmol min(-1) mg(-1), respectively. Due to its ability on decarboxylation of α-ketoisovalerate and α-ketoisocaproate, the recombinant E. coli strain showed potential application on isoamyl alcohol and isobutanol production.
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Xiaoyan
X
Chen
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Jingliang
J
Xu
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Liu
L
Yang
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China. yuanzh@ms.giec.ac.cn.
Zhenhong
Z
Yuan
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Shiyuan
S
Xiao
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Yu
Y
Zhang
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Cuiyi
C
Liang
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Minchao
M
He
Key Laboratory of Renewable Energy and Gas Hydrate, Chinese Academy of Sciences, Guangzhou, 510640, China.
Ying
Y
Guo
D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide) / Q000235:genetics / Q000378:metabolism*; D000426:Alcohol Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D000438:Alcohols / Q000737:chemistry* / Q000378:metabolism*; D000440:Butanols / Q000378:metabolism; D003653:Decarboxylation; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D005285:Fermentation / Q000187:drug effects; D005947:Glucose / Q000378:metabolism / Q000494:pharmacology; D045782:Hemiterpenes; D007651:Keto Acids / Q000378:metabolism / Q000494:pharmacology; D013294:Lactococcus lactis / Q000201:enzymology / Q000235:genetics; D060847:Metabolic Engineering; D000439:Pentanols / Q000378:metabolism; D013696:Temperature; D017355:Zymomonas / Q000201:enzymology / Q000235:genetics
D000438:Alcohols; D000440:Butanols; D045782:Hemiterpenes; D007651:Keto Acids; D000439:Pentanols; C040507:isobutyl alcohol; C001505:alpha-ketoisovalerate; C013082:alpha-ketoisocaproic acid; C029683:isopentyl alcohol; D000426:Alcohol Dehydrogenase; D042942:3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide); D005947:Glucose
3-methyl-1-butanol; Alcohol dehydrogenase; Isobutanol; α-keto acid decarboxylase