Terpenoids are natural products known for their medicinal and commercial applications. Metabolic engineering of microbial hosts for the production of valuable compounds, such as artemisinin and Taxol, has gained vast interest in the last few decades. The Generally Regarded As Safe (GRAS) 168 with its broad metabolic potential is considered one of these interesting microbial hosts. In the effort toward engineering as a cell factory for the production of the chemotherapeutic Taxol, we expressed the plant-derived taxadiene synthase (TXS) enzyme. TXS is responsible for the conversion of the precursor geranylgeranyl pyrophosphate (GGPP) to taxa-4,11-diene, which is the first committed intermediate in Taxol biosynthesis. Furthermore, overexpression of eight enzymes in the biosynthesis pathway was performed to increase the flux of the GGPP precursor. This was achieved by creating a synthetic operon harboring the genes encoding the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway (, , , , , , ) together with (encoding geranyl and farnesyl pyrophosphate synthases) responsible for providing farnesyl pyrophosphate (FPP). In addition, a vector harboring the gene (encoding geranylgeranyl pyrophosphate synthase, GGPPS, of ) to increase the supply of GGPP was introduced. The overexpression of the MEP pathway enzymes along with IspA and GGPPS caused an 83-fold increase in the amount of taxadiene produced compared to the strain only expressing TXS and relying on the innate pathway of . The total amount of taxadiene produced by that strain was 17.8 mg/l. This is the first account of the successful expression of taxadiene synthase in . We determined that the expression of GGPPS through the gene is essential for the formation of sufficient precursor, GGPP, in as its innate metabolism is not efficient in producing it. Finally, the extracellular localization of taxadiene production by overexpressing the complete MEP pathway along with IspA and GGPPS presents the prospect for further engineering aiming for semisynthesis of Taxol.
J Nat Prod. 1999 Jul;62(7):1068-71
[PMID:
10425147]
Proc Natl Acad Sci U S A. 2000 Jan 18;97(2):583-7
[PMID:
10639122]
J Nat Prod. 2000 Jan;63(1):37-40
[PMID:
10650075]
Arch Biochem Biophys. 2000 Jul 1;379(1):137-46
[PMID:
10864451]
Metab Eng. 2001 Jan;3(1):27-39
[PMID:
11162230]
Bioorg Med Chem. 2001 Sep;9(9):2237-42
[PMID:
11553461]
Proc Natl Acad Sci U S A. 2003 Apr 15;100(8):4678-83
[PMID:
12682299]
Biosci Biotechnol Biochem. 2004 Jan;68(1):132-7
[PMID:
14745175]
Cell Mol Life Sci. 2004 Jun;61(12):1401-26
[PMID:
15197467]
Biotechnol Bioeng. 2004 Oct 20;88(2):168-75
[PMID:
15449291]
Nat Rev Drug Discov. 2005 Mar;4(3):206-20
[PMID:
15729362]
Biotechnol Bioeng. 2006 Feb 5;93(2):212-24
[PMID:
16161138]
Biomol Eng. 2006 Dec;23(6):265-79
[PMID:
17049920]
Appl Microbiol Biotechnol. 2007 Feb;74(1):131-9
[PMID:
17115209]
Appl Microbiol Biotechnol. 2007 Jan;73(5):980-90
[PMID:
17115212]
Appl Microbiol Biotechnol. 2007 Jul;75(6):1377-84
[PMID:
17458547]
Trends Biotechnol. 2007 Sep;25(9):417-24
[PMID:
17681626]
Proc Natl Acad Sci U S A. 2008 May 6;105(18):6725-9
[PMID:
18436644]
Metab Eng. 2008 May-Jul;10(3-4):201-6
[PMID:
18485776]
Org Lett. 2008 Aug 21;10(16):3551-4
[PMID:
18630919]
Nat Prod Rep. 2008 Aug;25(4):656-61
[PMID:
18663389]
Microbiology. 2009 Jun;155(Pt 6):1758-75
[PMID:
19383706]
Biotechnol Lett. 2009 Nov;31(11):1789-93
[PMID:
19618272]
Curr Opin Drug Discov Devel. 1998 Nov;1(3):329-37
[PMID:
19649807]
Appl Microbiol Biotechnol. 2009 Oct;84(6):1003-19
[PMID:
19669755]
Phytochem Rev. 2006 Feb;5(1):75-97
[PMID:
20622989]
Science. 2010 Oct 1;330(6000):70-4
[PMID:
20929806]
Nat Protoc. 2011 Feb;6(2):242-51
[PMID:
21293463]
Biosci Biotechnol Biochem. 2011;75(12):2376-83
[PMID:
22146731]
Appl Microbiol Biotechnol. 2012 Jan;93(1):331-41
[PMID:
22159605]
Biotechnol Bioeng. 2013 Sep;110(9):2556-61
[PMID:
23483530]
Appl Environ Microbiol. 2013 Oct;79(20):6481-90
[PMID:
23956387]
Nat Protoc. 2014 Aug;9(8):1980-96
[PMID:
25058645]
FEBS J. 2014 Nov;281(21):4906-20
[PMID:
25181035]
Appl Microbiol Biotechnol. 2015 Jul;99(14):5907-15
[PMID:
25851715]
FEMS Microbiol Rev. 2015 Nov;39(6):968-84
[PMID:
26109597]
Appl Microbiol Biotechnol. 2015 Nov;99(22):9395-406
[PMID:
26373726]
J Gen Appl Microbiol. 2015;61(4):124-31
[PMID:
26377132]
J Bacteriol. 2016 Oct 7;198(21):2925-2935
[PMID:
27528508]
Biotechnol J. 2017 Apr;12(4):
[PMID:
28217906]
Bioresour Technol. 2017 Oct;241:430-438
[PMID:
28599221]
Sci Rep. 2017 Nov 8;7(1):15058
[PMID:
29118374]
Synth Syst Biotechnol. 2017 Aug 30;2(3):167-175
[PMID:
29318197]
J Biochem. 1981 May;89(5):1581-7
[PMID:
6792191]
J Biochem. 1982 Nov;92(5):1527-37
[PMID:
6818223]
J Bacteriol. 1995 May;177(9):2403-7
[PMID:
7730271]
Nature. 1994 Feb 17;367(6464):630-4
[PMID:
7906395]
Planta Med. 1997 Aug;63(4):291-5
[PMID:
9270370]
Nature. 1997 Nov 20;390(6657):249-56
[PMID:
9384377]
Biochemistry. 1998 Sep 22;37(38):13411-20
[PMID:
9748348]
