Quinate Hydroxycinnamoyl Transferase Research Articles

Hydroxycinnamoyltransferases Involved in the Accumulation of Caffeic Acid Esters in Gametophytes and Sporophytes of Equisetum arvense.

Four hydroxycinnamoyltransferases from Equisetum arvense L. were studied that catalyze the formation of mono-O-caffeoyl-meso-tartrate, di-O-caffeoyl-meso-tartrate, 5-O-caffeoylshikimate (dactylifrate), and 5-O-caffeoylquinate (chlorogenate). The enzymes were classified as coenzyme A (CoA)-ester-dependent acyltransferases (EC 2.3.1), i.e. hydroxycinnamoyl-CoA:meso-tartrate hydroxycinnamoyltransferase (CTT), hydroxycinnamoyl-CoA:caf-feoyl-meso-tartrate hydroxycinnamoyltransferase (CCT), hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyltransferase (CST), and hydroxycinnamoyl-CoA:quinate hydroxycinnamoyltransferase. The CTT, CCT, and CST were partially purified and separated from E. arvense gametophytes by hydrophobic interaction chromatography on Fractogel TSK Butyl-650 followed by molecular exclusion on fast protein liquid chromatography-Superdex-75 with 87-, 62-, and 130- fold enrichments and 12, 8, and 11% yields, respectively. The enzyme activities obtained with caffeoyl-CoA were 95 (CTT), 74 (CCT), and 200 [mu]kat (CST) kg-1 protein. The apparent native relative molecular weight values were found to be approximately 45,000 (CTT), 52,000 (CCT), and 50,000 (CST). Each enzyme showed highest activities at pH 7.5, the CCT and CST in Tris-HCl (1.2 and 1.0 M) and the CTT in imidazole-HCl (1.25 M). Enzyme activities were stimulated more than 3-fold by 100 mM ascorbate. The apparent energies of activation (kilojoules mol-1) were calculated to be 56 (CTT), 69 (CST), and 76 (CCT). The enzymes accepted cinnamoyl-CoA and various hydroxycinnamoyl-CoAs. The time course of the transferase activities along with that of a fourth one, hydroxycinnamoyl-CoA:quinate hydroxycinnamoyltransferase, and the pattern of product accumulation were determined during a 1-year growth period of the E. arvense sporophytes.

Partial purification and characterization of hydroxycinnamoyl CoA: Transferases from apple and date fruits

Two hydroxycinnamoyl CoA: transferases (EC 2.3.1.-) (HT) specific for shikimic and quinic acids, which catalyse the formation of 5- O-hydroxycinnamoyl-shikimic and/or -quinic esters, have been purified 82- and 470-fold from young green dates and apples, respectively. They showed highest activity with p-coumaroyl CoA compared with caffeoyl CoA and feruloyl CoA. Both enzymes were active with shikimic and quinic acids, but the relative ratios of the V max/ K m values for shikimic and quinic acids were found to be 100: 6 for date HT and 23: 100 for apple HT. We, therefore, term the two enzymes hydroxycinnamoyl CoA: shikimate hydroxycinnamoyl transferase (HST) for date and hydroxycinnamoyl CoA: quinate hydroxycinnamoyl transferase (HQT) for apple. The two HTs had similar M r and optimum pHs and K m values for cinnamoyl-CoA thioesters. The two enzymes exhibited distinct isoelectric point values: 4.63 for HST and 5.40 for HQT.

ヨウサイ (<i>Ipomoea aquatica</i> Forsk.) の低温障害発生に伴うK<sup>+</sup>漏出速度, 遊離アミノ酸含量並びにフェニルプロパノイド代謝の変化

Water convolvulus (Ipomoea aquatica Forsk.) is widely distributed as leafy vegetables in South-East Asia including southern area in Japan. It shows chilling injury manifested as browning of young leaves (2-3 immature leaves located at top part of each vine) and young stems (top part of each vine) when stored at temperatures about below 9°C. In the present study, the changes of K+ leakage, free amino acid contents, phenylpropanoid contents, and some enzyme activities relating to the biosynthesis of phenylpropanoids were inveatigated with reference to the chilling injury. The materials were sealed in polyethylene bags (0.03mm thickness) and stored at 1, 6, 9, 12, 15, 20, and 30°C.A break point of Arrhenius plot for K+leakage in young and mature leaves was found between 10 and 16°C, whereas the matured leaves showed only a little chilling-sensitivity. A marked increase of the rate of K+ leakage occurred in young leaves stored at 1°C, preceding the appearance of browning.As the contents of phenylpropanoids such as t-cinnamic, p-coumaric, caffeic and chlorogenic acids were investigated, they increased before the occurrence of chilling injury and then tended to decline almost simultaneously with the development of chilling injury. They also increased at 15°C, but the rise delayed and the extent was considerably lower than that at 1°C.Among the enzymes relating to the biosynthesis of chlorogenic acid, a main substrate of browning, hydroxycinnamoyl CoA ligase (CL) activity increased in both of 1°C and 15°C storages, then the increase was faster at 1°C reaching to a peak before the occurrence of chilling injury. Hydroxycinnamoyl CoA: quinate hydroxycinnamoyl transferase (CQT) activity showed a markedly higher increase in the 1°C storage.On the results, we present a conclusion of the low temperature induced browning mechanism as follows; exposure to low temperature leads to a rise of phenolic contents through activation of the enzymes such as CL and CQT. The accumulated phenolics induce the browning after its enzymic oxidation.

Purification and characterization of hydroxycinnamoyl D-glucose. Quinate hydroxycinnamoyl transferase in the root of sweet potato, Ipomoea batatas Lam.

We have previously proposed a chlorogenic acid biosynthetic pathway which involves a transesterification reaction between hydroxycinnamoyl D-glucose and D-quinic acid. The proposed pathway was based on tracer experimental results (Kojima, M., and Uritani, I. (1972) Plant Cell Physiol. 13, 311-319). The enzyme that catalyzes the above reaction has been purified 160-fold from sweet potato root (Ipomoea batatas Lam.) and characterized. The purified enzyme yielded one band of 26,000 daltons on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and its molecular weight was estimated to be 25,000 by gel filtration chromatography. Therefore, the enzyme seems to consist of a single polypeptide of 25,000-26,000 daltons. The isoelectric point of the enzyme was 8.6. The optimum pH of the enzyme reaction was 6.0. The enzyme did not require any metal for activity and showed a broad substrate specificity toward hydroxycinnamoyl D-glucose as donors. The Km and Vmax values were 3.7 mM and 8.5 units/mg of protein for t-cinnamoyl D-glucose, 3.9 mM and 15.1 units/mg of protein for p-coumaroyl D-glucose, and 14.3 mM and 38.1 units/mg of protein for caffeoyl D-glucose. The enzyme showed a strict substrate specificity toward D-quinic acid-related compounds as acceptors; the Km and Vmax values were 16.7 mM and 15.1 units/mg of protein for D-quinic acid, 250 mM and 19.0 units/mg of protein for shikimic acid, and there was no activity with either L-malic acid or meso-tartaric acid. The enzyme activity changed in a manner suggesting its involvement in chlorogenic acid biosynthesis during incubation of sliced sweet potato root tissues.

Orientation nouvelle du métabolisme des acides hydroxycinnamiques dans les fruits de tomates blessés (Lycopersicon esculentum)

A change in the metabolism of hydroxycinnamic acids in wounded tomato fruits (Lycopersicon esculentum).Healing of lesions in tomato fruits (Lycopersicon esculentum Mill. var. cerasiforme) is partly due to lignification of cells bordering the wounded zones. The pericarp of healthy fruits contains a high level of hydroxycinnamic derivatives but never shows lignification. Thus, the reaction of the fruit to wounding seems to be a change in the metabolism, leading to the formation of monomeric units of which lignins are constituted. Hydroxycinnamate:CoA ligase (EC 6.2.1.12; CL) and O‐methyltransferase (EC 2.1.1.6; OMT) appear to play an important role in this change. In wounded fruits CL acts preferentially on p‐coumarate and ferulate as compared to caffeate, and OMT is particularly active with 5‐OH ferulate as substrate. These changes lead to the formation of p‐coumaroyl CoA, feruloyl CoA and sinapate, which are incorporated into lignin. Phenylalanine ammonialyase (EC 4.3.1.5) and glucosyltransferase activities increase greatly after wounding, whereas the activity of hydroxycinnamoyl‐CoA:quinate hydroxycinnamoyl transferase decreases. These data complement those previously reported on peroxidases and suggest that, after the increase of enzyme activities, monomeric units are formed and then polymerized by some peroxidases specific for lignification.

Partial purification and properties of hydroxycinnamoyl-CoA: quinate hydroxycinnamoyl transferase from higher plants

The partial purification and characterization of hydroxycinnamoyl-CoA: quinate hydroxycinnamoyl transferase (CQT) from two plant sources growing as cell cultures are reported. The enzymes have been purified 50-and 16-fold, respectively, and show an absolute specificity for p-coumaroyl-CoA and caffeoyl-CoA as well as for quinate, and are responsible for the synthesis of p-coumaroylquinate and caffeoylquinate (chlorogenic acid). The distribution of this transferase activity in a variety of plant cell cultures and differentiated plants is reported.

Purification and properties of hydroxycinnamoyl CoA quinate hydroxycinnamoyl transferase from potatoes

A rapid method for the purification of hydroxycinnamoyl CoA quinate hydroxycinnamoyl transferase (CQT) from potato tubers which had been stored at low temperatures is described. The method involves affinity chromatography on Blue Sepharose with biospecific desorption of CQT with its substrate, CoA. Elution of the Blue Sepharose column with a gradient of CoA leads to the resolution of CQT, a protein with MW of ca 41500, into 3 peaks of activity; the largest peak elutes first. This fraction is purified × 1440 and gives a single band of protein after PAGE which suggests a high degree of purity. The properties of the 3 fractions of CQT, with respect to substrates and to a number of inhibitors, are described. The first and last eluting CQT fractions are specific for quinate and show no activity towards shikimate. The second peak, however, shows a small activity towards shikimate but this is thought to be due to an underlying peak of a shikimate specific enzyme. The major peak of CQT activity found in potatoes stored at 0° is absent from those stored at 10° throughout the period after harvest.

Induction by light of hydroxycinnamoyl-CoA: Quinate hydroxycinnamoyl transferase in buckwheat (Fagopyrum esculentum Moench): Absence of feed-forward control by trans-cinnamate

The light-induced increase in the activity of hydroxycinnamoyl-CoA: quinate hydroxycinnamoyl transferase (CQT) in hypocotyls of etiolated buckwheat seedlings is not suppressed by aminooxy acetic acid, an inhibitor of cinnamic acid formation in this tissue. Incubation of hypocotyls in 2 mmol l(-1) trans-cinnamate in the dark does not increase CQT activity. Thus, trans-cinnamate does not appear to mediate the effect of light on CQT in buckwheat hypocotyls.

Metabolisme des composes phenoliques chez le petunia VI. Intervention des chloroplastes dans la biosynthese de la naringenine et de l'acide chlorogenique

Enzymatic extract from Petunia leaves catalyze different reactions involved in the synthesis of hydroxycinnamoyl esters and flavonoids. Attempts to localize the individual enzymes within isolated chloroplasts show that these organelles: contain hydroxycinnamoyl-CoA—quinate—hydroxycinnamoyl transferase and flovanone-synthetase involved respectively in the biosynthesis of chlorogenic acid and naringenin; catalyze the isomerisation of chalcone to flavanone; are not able to activate the C 6—C 3 units to their CoA esters. These results are confirmed by feeding experiments on isolated chloroplasts using quinate or acetate 14C in combination (or not) with exogenously supplied “cinnamoyl-CoA”. These data seem to indicate that the activated cinnamic units, involved in the formation of chloroplastic phenols, are synthetized in other cellular compartment(s).

Trans-Cinnamic acid as a mediator of the light-stimulated increase in hydroxycinnamoyl-CoA: quinate hydroxycinnamoyl transferase.

trans-Cinnamic acid as a mediator of the light-stimulated increase in hydroxycinnamoyl-CoA: quinate hydroxycinnamoyl transferase.