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Anal Biochem. 2002 Mar 15;302(2):305-12.

Enzymatic synthesis and purification of aromatic coenzyme a esters.

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  • 1Department of Molecular, Cellular and Developmental Biology, University of Michigan, 830 North University Street, Ann Arbor, Michigan 48109-1048, USA.

Abstract

Two recombinant His-tagged proteins, a plant 4-coumarate:coenzyme A ligase (EC 6.2.1.12) and a bacterial benzoate:coenzyme A ligase (EC 6.2.1.25), were expressed in Escherichia coli and purified in a single step using Ni-chelating chromatography. Purified enzymes were used to synthesize cinnamoyl-coenzyme A (CoA), p-coumaroyl-CoA, feruloyl-CoA, caffeoyl-CoA, and benzoyl-CoA. Conversions up to 95% were achieved. Using a rapid solid-phase extraction procedure, the target CoA esters were isolated with yields of up to 80%. Structures were confirmed by analytical comparison with chemically synthesized reference compounds and electrospray ionization-mass spectrometry. The recombinant enzymes were stable for several months at -80 degrees C, thus providing a reliable and facile method to produce these delicate biological intermediates.

(C)2002 Elsevier Science (USA).

PMID:
11878812
[PubMed - indexed for MEDLINE]
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