title: Production of 7-O-methyl aromadendrin, a medicinally valuable flavonoid, in Escherichia coli.
abstract: 7-O-Methyl aromadendrin (7-OMA) is an aglycone moiety of one of the important flavonoid-glycosides found in several plants, such as Populus alba and Eucalyptus maculata, with various medicinal applications. To produce such valuable natural flavonoids in large quantity, an Escherichia coli cell factory has been developed to employ various plant biosynthetic pathways. Here, we report the generation of 7-OMA from its precursor, p-coumaric acid, in E. coli for the first time. Primarily, naringenin (NRN) (flavanone) synthesis was achieved by feeding p-coumaric acid and reconstructing the plant biosynthetic pathway by introducing the following structural genes: 4-coumarate-coenzyme A (CoA) ligase from Petroselinum crispum, chalcone synthase from Petunia hybrida, and chalcone isomerase from Medicago sativa. In order to increase the availability of malonyl-CoA, a critical precursor of 7-OMA, genes for the acyl-CoA carboxylase α and β subunits (nfa9890 and nfa9940), biotin ligase (nfa9950), and acetyl-CoA synthetase (nfa3550) from Nocardia farcinica were also introduced. Thus, produced NRN was hydroxylated at position 3 by flavanone-3-hydroxylase from Arabidopsis thaliana, which was further methylated at position 7 to produce 7-OMA in the presence of 7-O-methyltransferase from Streptomyces avermitilis. Dihydrokaempferol (DHK) (aromadendrin) and sakuranetin (SKN) were produced as intermediate products. Overexpression of the genes for flavanone biosynthesis and modification pathways, along with malonyl-CoA overproduction in E. coli, produced 2.7 mg/liter (8.9 μM) 7-OMA upon supplementation with 500 μM p-coumaric acid in 24 h, whereas the strain expressing only the flavanone modification enzymes yielded 30 mg/liter (99.2 μM) 7-OMA from 500 μM NRN in 24 h.
authors.affiliation: Laboratory of Molecular Biotechnology and Biomaterials, School of Chemical and Biological Engineering, Seoul National University, Seoul, South Korea.
authors.forename: Sailesh
authors.initials: S
authors.lastname: Malla
authors.forename: Mattheos A G
authors.initials: MA
authors.lastname: Koffas
authors.forename: Romas J
authors.initials: RJ
authors.lastname: Kazlauskas
authors.forename: Byung-Gee
authors.initials: BG
authors.lastname: Kim
mesh_terms: D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D053898:Biosynthetic Pathways / Q000235:genetics; D003373:Coumaric Acids / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005419:Flavonoids / Q000378:metabolism*; D000455:Medicago sativa / Q000201:enzymology / Q000235:genetics; D060847:Metabolic Engineering; D009615:Nocardia / Q000201:enzymology / Q000235:genetics; D028528:Petroselinum / Q000201:enzymology / Q000235:genetics; D032306:Petunia / Q000201:enzymology / Q000235:genetics; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D011422:Propionates; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D003373:Coumaric Acids; D005419:Flavonoids; D010940:Plant Proteins; D011422:Propionates; D011994:Recombinant Proteins; C080220:aromadedrin; C495469:p-coumaric acid
references.citation: Biol Pharm Bull. 2010;33(9):1494-9
references.pmid: 20823563
references.citation: Annu Rev Nutr. 2002;22:19-34
references.pmid: 12055336
references.citation: J Agric Food Chem. 2006 Feb 8;54(3):823-8
references.pmid: 16448189
references.citation: Appl Environ Microbiol. 2007 Jun;73(12):3877-86
references.pmid: 17468269
references.citation: Eur J Biochem. 1975 Aug 1;56(1):205-13
references.pmid: 240705
references.citation: J Ethnopharmacol. 1996 Jan;50(1):27-34
references.pmid: 8778504
references.citation: Appl Environ Microbiol. 2005 Sep;71(9):5610-3
references.pmid: 16151160
references.citation: Appl Environ Microbiol. 2003 May;69(5):2699-706
references.pmid: 12732539
references.citation: Drug Metab Dispos. 2006 Oct;34(10):1786-92
references.pmid: 16868069
references.citation: J Bacteriol. 1980 Jul;143(1):302-6
references.pmid: 6995433
references.citation: Biotechnol J. 2007 Oct;2(10):1214-34
references.pmid: 17935117
references.citation: Phytochemistry. 2000 Nov;55(6):481-504
references.pmid: 11130659
references.citation: Mol Pharm. 2008 Mar-Apr;5(2):257-65
references.pmid: 18333619
references.citation: Biotechnol Adv. 2008 Nov-Dec;26(6):548-60
references.pmid: 18682287
references.citation: Microbiol Mol Biol Rev. 2001 Dec;65(4):523-69, table of contents
references.pmid: 11729263
references.citation: Appl Microbiol Biotechnol. 2006 Mar;70(1):85-91
references.pmid: 16025328
references.citation: N Engl J Med. 2006 Dec 7;355(23):2427-43
references.pmid: 17145742
references.citation: J Nat Prod. 1999 Apr;62(4):601-4
references.pmid: 10217718
references.citation: Appl Microbiol Biotechnol. 2005 Sep;68(4):498-504
references.pmid: 15770480
references.citation: J Ethnopharmacol. 2008 Jul 23;118(2):257-63
references.pmid: 18508214
references.citation: JAMA. 2007 Sep 12;298(10):1189-95
references.pmid: 17848653
references.citation: J Bacteriol. 1983 Jul;155(1):113-21
references.pmid: 6345502
references.citation: J Gen Microbiol. 1988 Aug;134(8):2249-53
references.pmid: 3075658
references.citation: Diabetes Care. 2004 Jul;27(7):1741-2
references.pmid: 15220256
references.citation: Arch Pharm Res. 2006 Dec;29(12):1102-8
references.pmid: 17225458
references.citation: J Mass Spectrom. 2007 Sep;42(9):1136-47
references.pmid: 17565713
references.citation: Trends Plant Sci. 2000 Sep;5(9):380-6
references.pmid: 10973093
references.citation: Appl Environ Microbiol. 2009 Sep;75(18):5831-9
references.pmid: 19633125
references.citation: Metab Eng. 2006 Mar;8(2):172-81
references.pmid: 16384722
references.citation: BMJ. 2011 Mar 17;342:d1309
references.pmid: 21415101
references.citation: Pharmacol Rev. 2000 Dec;52(4):673-751
references.pmid: 11121513
references.citation: Pharmazie. 2000 Aug;55(8):623-4
references.pmid: 10989845
references.citation: Biochim Biophys Acta. 1973 Apr 12;302(2):457-64
references.pmid: 4699252
references.citation: Eur J Biochem. 1975 Jan 2;50(2):383-9
references.pmid: 236184
references.citation: Planta Med. 1982 Oct;46(2):124-5
references.pmid: 17396957