Document Detail


Key amino acid residues required for aryl migration catalysed by the cytochrome P450 2-hydroxyisoflavanone synthase.
MedLine Citation:
PMID:  12207646     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Isoflavonoids are distributed predominantly in leguminous plants, and play pivotal roles in the interaction of host plants with biological environments. Isoflavones in the diet also have beneficial effects on human health as phytoestrogens. The isoflavonoid skeleton is constructed by the CYP93C subfamily of cytochrome P450s in plant cells. The reaction consists of hydroxylation of the flavanone molecule at C-2 and an intramolecular 1,2-aryl migration from C-2 to C-3 to yield 2-hydroxyisoflavanone. In this study, with the aid of alignment of amino acid sequences of CYP93 family P450s and a computer-generated putative stereo structure of the protein, candidates for key amino acid residues in CYP93C2 responsible for the unique aryl migration in 2-hydroxyisoflavanone synthase reaction were identified. Microsomes of recombinant yeast cells expressing mutant proteins of CYP93C2 were prepared, and their catalytic activities tested. The reaction with the mutant in which Ser 310 in the centre of the I-helix was converted to Thr yielded increased formation of 3-hydroxyflavanone, a by-product of the 2-hydroxyisoflavanone synthase reaction, in addition to the major isoflavonoid product. More dramatically, the mutant in which Lys 375 in the end of beta-sheet 1-4 was replaced with Thr produced only 3-hydroxyflavanone and did not yield the isoflavonoid any longer. The roles of these amino acid residues in the catalysis and evolution of isoflavonoid biosynthesis are discussed.
Authors:
Yuji Sawada; Kengo Kinoshita; Tomoyoshi Akashi; Toshio Aoki; Shin-Ichi Ayabe
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Publication Detail:
Type:  Comparative Study; Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  The Plant journal : for cell and molecular biology     Volume:  31     ISSN:  0960-7412     ISO Abbreviation:  Plant J.     Publication Date:  2002 Sep 
Date Detail:
Created Date:  2002-09-04     Completed Date:  2002-12-09     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  9207397     Medline TA:  Plant J     Country:  England    
Other Details:
Languages:  eng     Pagination:  555-64     Citation Subset:  IM    
Affiliation:
Department of Applied Biological Sciences, Nihon University, Fujisawa, Kanagawa 252-8510, Japan.
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Amino Acids / metabolism*
Catalysis
Chromatography, High Pressure Liquid
Cytochrome P-450 Enzyme System / chemistry,  genetics,  metabolism*
Flavanones*
Flavonoids / chemistry,  metabolism
Gene Expression Regulation, Enzymologic
Hydrogen-Ion Concentration
Isoflavones / metabolism
Kinetics
Microsomes / metabolism
Mixed Function Oxygenases / metabolism
Molecular Sequence Data
Mutagenesis, Site-Directed
Mutation
Protein Binding
Protein Conformation
Yeasts / genetics,  metabolism
Chemical
Reg. No./Substance:
0/2,7,4'-trihydroxyisoflavanone; 0/3-hydroxyflavanone; 0/Amino Acids; 0/Flavanones; 0/Flavonoids; 0/Isoflavones; 0/cytochrome P-450 CYP93C2 (Glycyrrhiza echinata); 578-86-9/liquiritigenin; 9035-51-2/Cytochrome P-450 Enzyme System; EC 1.-/Mixed Function Oxygenases; EC 1.14.-/cytochrome P-450 CYP93B1 (Glycyrrhiza echinata)

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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