Identification and functional analysis of 2-hydroxyflavanone C-glucosyltransferase in soybean (Glycine max)

•2-Hydroxyflavanone C-glucosyltransferase (UGT708D1) was identified in soybean.•The catalytic residues were estimated on the basis of structural modeling.•Three conserved residues were shown to be necessary for C-glucosylation activity.•UGT708D1 with a point mutation (H20A) showed O-glucosylation ac...

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Bibliographic Details
Published in:FEBS letters 2015-07, Vol.589 (15), p.1778-1786
Main Authors: Hirade, Yoshihiro, Kotoku, Naoyuki, Terasaka, Kazuyoshi, Saijo-Hamano, Yumiko, Fukumoto, Akemi, Mizukami, Hajime
Format: Article
Language:English
Subjects:
2-Hydroxyflavanone C-glucosyltransferase
Amino Acid Sequence
C-Glucoside biosynthesis
C-glucosyltransferase
Catalysis
Catalytic Domain
Catalytic residues
CGT
Chromatography, High Pressure Liquid
EAS
electrophilic aromatic substitution
Enzymatic aryl-C-glucosylation
EST
expressed sequence tag
Glucosyltransferases - chemistry
Glucosyltransferases - classification
Glucosyltransferases - metabolism
Glycine max - enzymology
high-performance liquid chromatography
HPLC
Mass Spectrometry
Molecular Sequence Data
O-glucosyltransferase
OGT
Oryza sativa C-glucosyltransferase
OsCGT
Phylogeny
plant secondary product glycosyltransferase
PSPG
Recombinant Proteins - chemistry
Recombinant Proteins - metabolism
Secondary metabolites
Sequence Homology, Amino Acid
Structural modeling
UDP
UDP-sugar-dependent glycosyltransferase
UGT
uridine diphosphate
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Description
Summary:•2-Hydroxyflavanone C-glucosyltransferase (UGT708D1) was identified in soybean.•The catalytic residues were estimated on the basis of structural modeling.•Three conserved residues were shown to be necessary for C-glucosylation activity.•UGT708D1 with a point mutation (H20A) showed O-glucosylation activity. C-Glucosyltransferase is an enzyme that mediates carbon–carbon bond formation to generate C-glucoside metabolites. Although it has been identified in several plant species, the catalytic amino acid residues required for C-glucosylation activity remain obscure. Here, we identified a 2-hydroxyflavanone C-glucosyltransferase (UGT708D1) in soybean. We found that three residues, His20, Asp85, and Arg292, of UGT708D1 were located at the predicted active site and evolutionarily conserved. The substitution of Asp85 or Arg292 with alanine destroyed C-glucosyltransferase activity, whereas the substitution of His20 with alanine abolished C-glucosyltransferase activity but enabled O-glucosyltransferase activity. The catalytic mechanism is discussed on the basis of the findings.
ISSN:0014-5793
1873-3468
DOI:10.1016/j.febslet.2015.05.010