Functional characterization of two p-coumaroyl ester 3′-hydroxylase genes from coffee tree: evidence of a candidate for chlorogenic acid biosynthesis

Abstract

Chlorogenic acid (5-CQA) is one of the major soluble phenolic compounds that is accumulated in coffee green beans. With other hydroxycinnamoyl quinic acids (HQAs), this compound is accumulated in particular in green beans of the cultivated species Coffea canephora. Recent work has indicated that the biosynthesis of 5-CQA can be catalyzed by a cytochrome P450 enzyme, CYP98A3 from Arabidopsis. Two full-length cDNA clones (CYP98A35 and CYP98A36) that encode putative p-coumaroylester 3'-hydroxylases (C3'H) were isolated from C. canephora cDNA libraries. Recombinant protein expression in yeast showed that both metabolized p-coumaroyl shikimate at similar rates, but that only one hydroxylates the chlorogenic acid precursor p-coumaroyl quinate. CYP98A35 appears to be the first C3'H capable of metabolising p-coumaroyl quinate and p-coumaroyl shikimate with the same efficiency. We studied the expression patterns of both genes on 4-month old C. canephora plants and found higher transcript levels in young and in highly vascularized organs for both genes. Gene expression and HQA content seemed to be correlated in these organs. Histolocalization and immunolocalization studies revealed similar tissue localization for caffeoyl quinic acids and p-coumaroylester 3'-hydroxylases. The results indicated that HQA biosynthesis and accumulation occurred mainly in the shoot tip and in the phloem of the vascular bundles. The lack of correlation between gene expression and HQA content observed in some organs is discussed in terms of transport and accumulation mechanisms.

Novel Hydroxycinnamoyl-Coenzyme A Quinate Transferase Genes from Artichoke Are Involved in the Synthesis of Chlorogenic Acid

“…7; Mahesh et al, 2007). Our data suggest that the route of CGA synthesis in artichoke is likely modulated according to substrate availability, and probably in vivo, CGA synthesis occurs preferentially via route 1, involving HCT, C3#H, and HQT (Fig.…”

“…This is also suggested by the evidence that a recently isolated C3#H enzyme from artichoke proved to be more efficient, within route 1b, in the conversion of p-coumaroylshikimate into caffeoylshikimate . However, other possible isoforms of C3#H could be involved in route 2, as in coffee (Mahesh et al, 2007).…”

“…In the first, hydroxycinnamoyl-CoA quinate hydroxycinnamoyl transferase (HQT) catalyzes the formation of CGA from caffeoyl-CoA and quinic acid (route 1; Niggeweg et al, 2004). Caffeoyl-CoA is supplied by the combined activities of hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferase (HCT) and p-coumaroyl ester 3# hydroxylase (C3#H) via a p-coumaroylshikimate intermediate (route 1b; Mahesh et al, 2007). The second proposed route is based on the synthesis of p-coumaroylquinate by HCT, followed by hydroxylation by C3#H (Ulbrich andZenk, 1979, Hoffmann et al, 2003).…”

Isolation and mapping of a C3′H gene (CYP98A49) from globe artichoke, and its expression upon UV-C stress

“…Most of the members of the CYP98 family, described to date, metabolize shikimate esters of p-coumaric acid more efficiently than quinate esters (Schoch et al 2001(Schoch et al , 2006Morant et al 2007). On the other hand CYP98A35 from coffee is capable of metabolizing p-coumaroylquinate and p-coumaroylshikimate with the same efficiency (Mahesh et al 2007). CYP98A49 from globe artichoke appears to show a lower affinity for quinate esters than shikimate esters (Fig.…”

“…The 3 0 -hydroxylation of the phenylpropanoid ring is essential not only for the synthesis of lignin (Franke et al 2002;Abdulrazzak et al 2006;Chen and Dixon 2007), but is also an important step in the synthesis of chlorogenic acid (Schoch et al 2006;Mahesh et al 2007). The CYP98 family of plant cytochromes P450 has been designated as the family of enzyme that performs the meta-hydroxylation step in the phenylpropanoid pathway ( Fig.…”