Enzymatic synthesis and purification of caffeoyl-CoA, p-coumaroyl-CoA, and feruloyl-CoA.
An enzyme preparation from wheat seedlings containing p-coumaroyl:CoA ligase activity was used to synthesize caffeoyl-CoA, p-coumaroyl-CoA, and feruloyl-CoA. The same enzyme preparation also contains caffeic acid-3-O-methyl transferase and caffeoyl-CoA-3-O-methyl transferase activities. The maximum activity was found in enzyme preparation from 2-day-old seedlings, where 15-20% of the hydroxy cinnamic acid could be converted into the corresponding thioester. This yield is a result of an equilibrium between the ligase and a thioesterase also present in the crude enzyme preparation. The activity of caffeic acid 3-O-methyl transferase and caffeoyl-CoA 3-O-methyl transferase enables the production of (14)C-labeled feruloyl-CoA when using S-adenosyl-l-[methyl-(14)C]-methionine as methyl donor. The produced thioesters can be purified by reverse phase HPLC using a phosphoric acid-acetonitrile gradient.
Obel
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Plant Biochemistry Laboratory, Department of Plant Biology, The Royal Veterinary and Agricultural University, 40 Thorvaldsensvej, Frederiksberg C., Dk-1871, Denmark.
Scheller
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HV
D000097:Acetonitriles / Q000737:chemistry; D000214:Acyl Coenzyme A / Q000138:chemical synthesis* / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D003373:Coumaric Acids / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004950:Esterases / Q000378:metabolism; D004952:Esters / Q000138:chemical synthesis / Q000302:isolation & purification; D008025:Ligases / Q000378:metabolism; D010756:Phosphoric Acids / Q000737:chemistry; D012436:S-Adenosylmethionine / Q000737:chemistry; D013379:Substrate Specificity; D013997:Time Factors; D014908:Triticum / Q000201:enzymology
D000097:Acetonitriles; D000214:Acyl Coenzyme A; D003373:Coumaric Acids; D004952:Esters; D010756:Phosphoric Acids; C119167:feruloyl-CoA; C058644:4-coumaroyl-coenzyme A; C058645:caffeoyl-coenzyme A; D012436:S-Adenosylmethionine; C030242:phosphoric acid; D004950:Esterases; D008025:Ligases; C032159:acetonitrile