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Plant Sci. 2013 Oct;211:61-9. doi: 10.1016/j.plantsci.2013.07.003. Epub 2013 Jul 12.

Purification and characterization of a betanidin glucosyltransferase from Amaranthus tricolor L catalyzing non-specific biotransformation of flavonoids.

Author information

1
Plant Biotechnology Laboratory, Department of Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur, West Bengal, India.

Abstract

Betacyanins are the major pigments present in Amaranthus tricolor, a leafy vegetable consumed globally. The terminal glycosylation of the aglycone betanidin is an important step in the biosynthesis of this natural red antioxidant pigment. A betanidin 5-O-glucosyltransferase (BGT) was fully purified to 134 folds (specific activity, 265.2 nkat mg(-1)) from the red amaranth by ammonium sulfate precipitation followed by hydrophobic interaction, anion exchange and size exclusion chromatography. Homogeneity of the purified protein was confirmed by 2-dimensional polyacrylamide gel electrophoresis (2D PAGE). The molecular weight of the enzyme determined by liquid chromatography-mass spectrometry (LC-MS) was found to be 62.8 kDa. Furthermore, the enzyme glycosylated flavonoids (kaempferol and quercetin) but not anthocyanidins, presence of which is mutually exclusive to betacyanin accumulating plants. The apparent Km (344±2.34 μM) and Vmax (17.24 μM min(-1)) of the enzyme were determined by LC-MS/MS. Peptide mass fingerprinting of the purified protein showed 38.4% coverage of peptide masses with anthocyanidin 3-O-glucosyltransferase from Zea mays. Study on this purified enzyme, for the first time, revealed its role of glycosylation in biosynthesis of betacyanin in A. tricolor and indicates promiscuous substrate-specificity.

KEYWORDS:

Amaranthus tricolor L; BGT; Betanidin glucosyltransferase; Flavonoids; GT; LC–MS; Liquid chromatography–mass spectrometry; PMF; Peptide mass fingerprinting; betanidin glucosyltransferase; glycosyltransferase; liquid chromatography–mass spectrometry; peptide mass fingerprinting

PMID:
23987812
DOI:
10.1016/j.plantsci.2013.07.003
[Indexed for MEDLINE]

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