Boosting the free fatty acid synthesis of Escherichia coli by expression of a cytosolic Acinetobacter baylyi thioesterase
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- 作者：Yanning Zheng (1) (2)
Lingling Li (1) (3)
Qiang Liu (1) (3)
Wen Qin (3)
Jianming Yang (1) (2)
Yujin Cao (1)
Xinglin Jiang (1) (2)
Guang Zhao (1)
Mo Xian (1) - 关键词：Thioesterase ; Acinetobacter baylyi ; Escherichia coli ; Free fatty acid ; Substrate specificity ; Active ; site residues
- 刊名：Biotechnology for Biofuels
- 出版年：2012
- 出版时间：December 2012
- 年：2012
- 卷：5
- 期：1
- 全文大小：500KB
- 参考文献：1. Chasan R: Engineering fatty acids-The long and short of it. / Plant Cell 1995, 7:235-37.
2. Gao Q, Wang W, Zhao H, Lu X: Effects of fatty acid activation on photosynthetic production of fatty acid-based biofuels in Synechocystis sp. PCC6803. / Biotechnol Biofuels 2012, 5:17. CrossRef
3. Pua F-l, Fang Z, Zakaria S, Guo F, Chia C-h: Direct production of biodiesel from high-acid value Jatropha oil with solid acid catalyst derived from lignin. / Biotechnol Biofuels 2011, 4:56. CrossRef
4. Steen EJ, Kang Y, Bokinsky G, Hu Z, Schirmer A, McClure A, del Cardayre SB, Keasling JD: Microbial production of fatty-acid-derived fuels and chemicals from plant biomass. / Nature 2010, 463:559-62. CrossRef
5. Schirmer A, Rude MA, Li X, Popova E, del Cardayre SB: Microbial biosynthesis of alkanes. / Science 2010, 329:559-62. CrossRef
6. Dehesh K, Edwards P, Hayes T, Cranmer AM, Fillatti J: Two novel thioesterases are key determinants of the bimodal distribution of acyl chain length of Cuphea palustvis seed oil. / Plant Physiol 1996, 110:203-10. CrossRef
7. Rehm BHA, Steinbüchel A: Heterologous expression of the acyl-acyl carrier protein thioesterase gene from the plant Umbellularia californica mediates polyhydroxyalkanoate biosynthesis in recombinant Escherichia coli . / Appl Microbiol Biotechnol 2001, 55:205-09. CrossRef
8. Voelker TA, Davies HM: Alteration of the specificity and regulation of fatty acid synthesis of Escherichia coli by expression of a plant medium-chain acyl-acyl carrier protein thioesterase. / J Bacteriol 1994, 176:7320-327.
9. Voelker TA, Worrell AC, Anderson L, Bleibaum J, Fan C, Hawkins DJ, Radke SE, Davies HM: Fatty acid biosynthesis redirected to medium chains in transgenic oilseed plants. / Science 1992, 257:72-4. CrossRef
10. Zhang X, Li M, Agrawal A, San K-Y: Efficient free fatty acid production in Escherichia coli using plant acyl-ACP thioesterases. / Metab Eng 2011, 13:713-22. CrossRef
11. Cao Y, Yang J, Xian M, Xu X, Liu W: Increasing unsaturated fatty acid contents in Escherichia coli by coexpression of three different genes. / Appl Microbiol Biotechnol 2010, 87:271-80. CrossRef
12. Hoover S, Marner W, Brownson A, Lennen R, Wittkopp T, Yoshitani J, Zulkifly S, Graham L, Chaston S, McMahon K, Pfleger B: Bacterial production of free fatty acids from freshwater macroalgal cellulose. / Appl Microbiol Biotechnol 2011, 91:435-46. CrossRef
13. Liu T, Vora H, Khosla C: Quantitative analysis and engineering of fatty acid biosynthesis in E. coli. / Metab Eng 2010, 12:378-86. CrossRef
14. Liu X, Sheng J, Curtiss R: Fatty acid production in genetically modified cyanobacteria. / Proc Natl Acad Sci USA 2011, 108:6899-904. CrossRef
15. Lu X, Vora H, Khosla C: Overproduction of free fatty acids in E. coli : Implications for biodiesel production. / Metab Eng 2008, 10:333-39. CrossRef
16. Yang J, Xian M, Su S, Zhao G, Nie Q, Jiang X, Zheng Y, Liu W: Enhancing production of bio-isoprene using hybrid MVA pathway and isoprene synthase in E. coli. / PLoS One 2012, 7:e33509. CrossRef
17. Jeon E, Lee S, Won J-I, Han SO, Kim J, Lee J: Development of Escherichia coli MG1655 strains to produce long chain fatty acids by engineering fatty acid synthesis (FAS) metabolism. / Enzyme Microb Tech 2011, 49:44-1. CrossRef
18. Reiser S, Somerville C: Isolation of mutants of Acinetobacter calcoaceticus deficient in wax ester synthesis and complementation of one mutation with a gene encoding a fatty acyl coenzyme A reductase. / J Bacteriol 1997, 179:2969-975.
19. Santala S, Efimova E, Kivinen V, Larjo A, Aho T, Karp M, Santala V: Improved triacylglycerol production in Acinetobacter baylyi ADP1 by metabolic engineering. / Microb Cell Fact 2011, 10:36. CrossRef
20. Bendtsen JD, Nielsen H, Heijne G, Brunak S: Improved prediction of signal peptides: SignalP 3.0. / J Mol Biol 2004, 340:783-95. CrossRef
21. Nielsen H, Engelbrecht J, Brunak S, Heijne G: Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. / Protein Eng 1997, 10:1-. CrossRef
22. Lee Y-L, Lee L-C, Leu R-J, Shaw J-F: Functional role of catalytic triad and oxyanion hole-forming residues on enzyme activity of Escherichia coli thioesterase I/protease I/phospholipase L1. / Biochem J 2006, 397:69. CrossRef
23. Lo Y-C, Lin S-C, Shaw J-F, Liaw Y-C: Crystal structure of Escherichia coli thioesterase I/protease I/lysophospholipase L1: consensus sequence blocks constitute the catalytic center of SGNH-hydrolases through a conserved hydrogen bond network. / J Mol Biol 2003, 330:539-51. CrossRef
24. Tyukhtenkoa SI, Litvinchuka AV, Changa C-F, Leub R-J, Shawb J-F, Huang T-H: NMR studies of the hydrogen bonds involving the catalytic triad of Escherichia coli thioesterase/protease I. / FEBS Lett 2002, 528:203-06. CrossRef
25. Mayer KM, Shanklin J: Identification of amino acid residues involved in substrate specificity of plant acyl-ACP thioesterases using a bioinformatics-guided approach. / BMC Plant Biol 2007, 7:1. CrossRef
26. Pazirandeh M, Chirala SS, Wakil SJ: Site-directed mutagenesis studies on the recombinant thioesterase domain of chicken fatty acid synthase expressed in Escherichia coli . / J Biol Chem 1991, 266:20946-0952.
27. Witkowski A, Naggert J, Witkowska HE, Randhawa ZI, Smith S: Utilization of an active serine 101 ?cysteine mutant to demonstrate the proximity of the catalytic serine 101 and histidine 237 residues in thioesterase II. / J Biol Chem 1992, 267:18488-8492.
28. Yuan L, Nelson BA, Caryl G: The catalytic cysteine and histidine in the plant acyl-acyl carrier protein thioesterases. / J Biol Chem 1996, 271:3417-419. CrossRef
29. Cho H, Cronan JE: Defective export of a periplasmic enzyme disrupts regulation of fatty acid synthesis. / J Biol Chem 1995, 270:4216-219. CrossRef
30. Davis MS, Solbiati J, Cronan JE: Overproduction of acetyl-CoA carboxylase activity increases the rate of fatty acid biosynthesis in Escherichia coli . / J Biol Chem 2000, 275:28593-8598. CrossRef
31. Xu H-M, Zhang G-Y, Ji X-D, Cao L, Shu L, Hua Z-C: Expression of soluble, biologically active recombinant human endostatin in Escherichia coli . / Protein Expr Purif 2005, 41:252-58. CrossRef
32. Goh LL, Loke P, Singh M, Sim TS: Soluble expression of a functionally active Plasmodium falciparum falcipain-2 fused to maltose-binding protein in Escherichia coli . / Protein Expr Purif 2003, 32:194-01. CrossRef
33. Cao Y, Xian M, Yang J, Xu X, Liu W, Li L: Heterologous expression of stearoyl-acyl carrier protein desaturase (S-ACP-DES) from Arabidopsis thaliana in Escherichia coli . / Protein Expr Purif 2010, 69:209-14. CrossRef
34. Lennen RM, Braden DJ, West RM, Dumesic JA, Pfleger BF: A process for microbial hydrocarbon synthesis: Overproduction of fatty acids in Escherichia coli and catalytic conversion to alkanes. / Biotechnol Bioeng 2010, 106:193-02. CrossRef
35. Torija MJ, Beltran G, Novo M, Poblet M, Rozès N, Guillamón JM, Mas A: Effect of the nitrogen source on the fatty acid composition of Saccharomyces cerevisiae . / Food Microbiol 2003, 20:255-58. CrossRef
36. Meng X, Yang J, Cao Y, Li L, Jiang X, Xu X, Liu W, Xian M, Zhang Y: Increasing fatty acid production in E. coli by simulating the lipid accumulation of oleaginous microorganisms. / J Ind Microbiol Biotechnol 2011, 38:919-25. CrossRef
37. Guzman LM, Belin D, Carson MJ, Beckwith J: Tight regulation, modulation, and high-level expression by vectors containing the arabinose P BAD promoter. / J Bacteriol 1995, 177:4121-130.
38. Hemsley A, Arnheim N, Toney MD, Cortopassi G, Galas DJ: A simple method for site-directed mutagenesis using the polymerase chain reaction. / Nucleic Acids Res 1989, 17:6545-551. CrossRef
39. Guan W, Zhao H, Lu X, Wang C, Yang M, Bai F: Quantitative analysis of fatty-acid-based biofuels produced by wild-type and genetically engineered cyanobacteria by gas chromatography–mass spectrometry. / J Chromatogr A 2011, 1218:8289-293. CrossRef - 作者单位：Yanning Zheng (1) (2)
Lingling Li (1) (3)
Qiang Liu (1) (3)
Wen Qin (3)
Jianming Yang (1) (2)
Yujin Cao (1)
Xinglin Jiang (1) (2)
Guang Zhao (1)
Mo Xian (1)
1. Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, 266101, China
2. University of Chinese Academy of Sciences, Beijing, 100049, China
3. College of Food Science, Sichuan Agricultural University, Yaan, 625014, China - ISSN：1754-6834
文摘
Background Thioesterases remove the fatty acyl moiety from the fatty acyl-acyl carrier proteins (ACPs), releasing them as free fatty acids (FFAs), which can be further used to produce a variety of fatty acid-based biofuels, such as biodiesel, fatty alcohols and alkanes. Thioesterases play a key role in the regulation of the fatty acid synthesis in Escherichia coli. Therefore, exploring more promising thioesterases will contribute to the development of industrial microbial lipids production. Results We cloned and expressed a cytosolic Acinetobacter baylyi thioesterase (‘AcTesA) in E. coli by deleting its leader sequence. Protein sequence alignment, structure modeling and site-directed mutagenesis demonstrated that Ser10, Gly48, Asn77, Asp158 and His161 residues composed the active centre of ‘AcTesA. The engineered strain that overexpressed ‘AcTesA achieved a FFAs titer of up to 501.2 mg/L in shake flask, in contrast to only 20.5 mg/L obtained in wild-type E. coli, demonstrating that the expression of ‘AcTesA indeed boosted the synthesis of FFAs. The ‘AcTesA exhibited a substrate preference towards the C8-C16 acyl groups, with C14:0, C16:1, C12:0 and C8:0 FFAs being the top four components. Optimization of expression level of ‘AcTesA made the FFAs production increase to 551.3 mg/L. The FFAs production further increased to 716.1 mg/L by optimization of the culture medium. Fed-batch fermentation was also carried out to evaluate the FFAs production in a scaleable process. Finally, 3.6 g/L FFAs were accumulated within 48 h, and a maximal FFAs yield of 6.1% was achieved in 12-6 h post induction. Conclusions For the first time, an A. baylyi thioesterase was cloned and solubly expressed in the cytosol of E. coli. This leaderless thioesterase (‘AcTesA) was found to be capable of enhancing the FFAs production of E. coli. Without detailed optimization of the strain and fermentation, the finally achieved 3.6 g/L FFAs is encouraging. In addition, ‘AcTesA exhibited different substrate specificity from other thioesterases previously reported, and can be used to supply the fatty acid-based biofuels with high quality of FFAs. Altogether, this study provides a promising thioesterase for FFAs production, and is of great importance in enriching the library of useful thioesterases.