Engineering the production of major catechins by Escherichia coli carrying metabolite genes of Camellia sinensis.
A mimicked biosynthetic pathway of catechin metabolite genes from C. sinensis, consisting of flavanone 3 hydroxylase (F3H), dihydroflavonol reductase (DFR), and leucoanthocyanidin reductase (LCR), was designed and arranged in two sets of constructs: (a) single promoter in front of F3H and ribosome-binding sequences both in front of DFR and LCR; (b) three different promoters with each in the front of the three genes and ribosome-binding sequences at appropriate positions. Recombinant E. coli BL (DE3) harbouring the constructs were cultivated for 65 h at 26 °C in M9 medium consisting of 40 g/L glucose, 1 mM IPTG, and 3 mM eriodictyol. Compounds produced were extracted in ethyl acetate in alkaline conditions after 1 h at room temperature and identified by HPLC. Two of the four major catechins, namely, (-)-epicatechin (0.01) and (-)-epicatechin gallate (0.36 mg/L), and two other types ((+)-catechin hydrate (0.13 mg/L) and (-)-catechin gallate (0.04 mg/L)) were successfully produced.
Department of Food Science, Faculty of Food Science and Technology, Universiti Putra Malaysia UPM 43400, Serdang Selangor, Malaysia.
Kabir Mustapha
KM
Umar
S M
SM
Abdulkarim
Son
S
Radu
Azizah
A
Abdul Hamid
Nazamid
N
Saari
D000429:Alcohol Oxidoreductases / Q000235:genetics; D000872:Anthocyanins / Q000378:metabolism; D028241:Camellia sinensis / Q000378:metabolism*; D002392:Catechin / Q000737:chemistry / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D017343:Genes, Plant / Q000235:genetics*; D005818:Genetic Engineering / Q000379:methods; D006899:Mixed Function Oxygenases / Q000235:genetics; D010088:Oxidoreductases / Q000235:genetics; D011401:Promoter Regions, Genetic; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000737:chemistry; D012270:Ribosomes / Q000737:chemistry; D013696:Temperature; D013997:Time Factors
D000872:Anthocyanins; D011994:Recombinant Proteins; D002392:Catechin; D006899:Mixed Function Oxygenases; D010088:Oxidoreductases; D000429:Alcohol Oxidoreductases; C056570:dihydroflavanol 4-reductase; C048797:flavanone 3-dioxygenase