Planta

, Volume 226, Issue 6, pp 1501–1509

Purification and characterization of UDP-glucose: anthocyanin 3′,5′-O-glucosyltransferase from Clitoria ternatea

  • Koichiro Kogawa
  • Naoki Kato
  • Kohei Kazuma
  • Naonobu Noda
  • Masahiko Suzuki
Original Article

Abstract

A UDP-glucose: anthocyanin 3′,5′-O-glucosyltransferase (UA3′5′GT) (EC 2.4.1.-) was purified from the petals of Clitoria ternatea L. (Phaseoleae), which accumulate polyacylated anthocyanins named ternatins. In the biosynthesis of ternatins, delphinidin 3-O-(6″-O-malonyl)-β-glucoside (1) is first converted to delphinidin 3-O-(6″-O-malonyl)-β-glucoside-3′-O-β-glucoside (2). Then 2 is converted to ternatin C5 (3), which is delphinidin 3-O-(6″-O-malonyl)-β-glucoside-3′,5′-di-O-β-glucoside. UA3′5′GT is responsible for these two steps by transferring two glucosyl groups in a stepwise manner. Its substrate specificity revealed the regioselectivity to the anthocyanin′s 3′- or 5′-OH groups. Its kinetic properties showed comparable kcat values for 1 and 2, suggesting the subequality of these anthocyanins as substrates. However, the apparent Km value for 1 (3.89 × 10−5 M), which is lower than that for 2 (1.38 × 10−4 M), renders the kcat/Km value for 1 smaller, making 1 catalytically more efficient than 2. Although the apparent Km value for UDP-glucose (6.18 × 10−3 M) with saturated 2 is larger than that for UDP-glucose (1.49 × 10−3 M) with saturated 1, the kcat values are almost the same, suggesting the UDP-glucose binding inhibition by 2 as a product. UA3′5′GT turns the product 2 into a substrate possibly by reversing the B-ring of 2 along the C2-C1′ single bond axis so that the 5′-OH group of 2 can point toward the catalytic center.

Keywords

Anthocyanin 3′-O-glucosyltransferase Anthocyanin 3′,5′-O-glucosyltransferase Anthocyanin 5′-O-glucosyltransferase Anthocyanidin glycoside biosynthesis Clitoria Polyacylated anthocyanins 

Abbreviations

DTT

Dithiothreitol

pAPMSF

p-Amidinophenylmethylsulfonylfluoride

TFA

Trifluoroacetic acid

UA3′GT

UDP-glucose: anthocyanin 3′-O-glucosyltransferase

UA3GT

UDP-glucose: anthocyanidin 3-O-glucosyltransferase

UA3′5′GT

UDP-glucose: anthocyanin 3′,5′-O-glucosyltransferase

UA5′GT

UDP-glucose: anthocyanin 5′-O-glucosyltransferase

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Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Koichiro Kogawa
    • 1
    • 2
  • Naoki Kato
    • 1
    • 3
  • Kohei Kazuma
    • 1
  • Naonobu Noda
    • 1
    • 4
  • Masahiko Suzuki
    • 1
    • 5
  1. 1.Division of Cell EngineeringAomori Green BioCentreAomori, AomoriJapan
  2. 2.Kamikita Agriculture, Forestry, and Fisheries Office, Misawa BranchMisawa, Aomori Japan
  3. 3.Aomori Ornamental Experiment StationGoshizawa, Aomori, AomoriJapan
  4. 4.National Institute of Floricultural Sciences Tsukuba, IbarakiJapan
  5. 5.Department of Horticulture, Graduate School of AgricultureHokkaido UniversitySapporoJapan

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