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Plant Physiol

Title:		"Cloning of beta-primeverosidase from tea leaves, a key enzyme in tea aroma formation"
Author(s):		Mizutani M; Nakanishi H; Ema J; Ma SJ; Noguchi E; Inohara-Ochiai M; Fukuchi-Mizutani M; Nakao M; Sakata K;
Address:		"Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan. mizutani@scl.kyoto-u.ac.jp"
Journal Title:		Plant Physiol
Year:		2002
Volume:		130
Issue:		4
Page Number:		2164 - 2176
DOI:		10.1104/pp.102.011023
ISSN/ISBN:		0032-0889 (Print) 1532-2548 (Electronic) 0032-0889 (Linking)
Abstract:		"A beta-primeverosidase from tea (Camellia sinensis) plants is a unique disaccharide-specific glycosidase, which hydrolyzes aroma precursors of beta-primeverosides (6-O-beta-D-xylopyranosyl-beta-D-glucopyranosides) to liberate various aroma compounds, and the enzyme is deeply concerned with the floral aroma formation in oolong tea and black tea during the manufacturing process. The beta-primeverosidase was purified from fresh leaves of a cultivar for green tea (C. sinensis var sinensis cv Yabukita), and its partial amino acid sequences were determined. The beta-primeverosidase cDNA has been isolated from a cDNA library of cv Yabukita using degenerate oligonucleotide primers. The cDNA insert encodes a polypeptide consisting of an N-terminal signal peptide of 28 amino acid residues and a 479-amino acid mature protein. The beta-primeverosidase protein sequence was 50% to 60% identical to beta-glucosidases from various plants and was classified in a family 1 glycosyl hydrolase. The mature form of the beta-primeverosidase expressed in Escherichia coli was able to hydrolyze beta-primeverosides to liberate a primeverose unit and aglycons, but did not act on 2-phenylethyl beta-D-glucopyranoside. These results indicate that the beta-primeverosidase selectively recognizes the beta-primeverosides as substrates and specifically hydrolyzes the beta-glycosidic bond between the disaccharide and the aglycons. The stereochemistry for enzymatic hydrolysis of 2-phenylethyl beta-primeveroside by the beta-primeverosidase was followed by (1)H-nuclear magnetic resonance spectroscopy, revealing that the enzyme hydrolyzes the beta-primeveroside by a retaining mechanism. The roles of the beta-primeverosidase in the defense mechanism in tea plants and the floral aroma formation during tea manufacturing process are also discussed"
Keywords:		"Amino Acid Sequence Base Sequence Biological Transport Cloning, Molecular DNA, Complementary/chemistry/genetics Escherichia coli/genetics/metabolism Gene Expression Regulation, Enzymologic Gene Expression Regulation, Plant Glycoside Hydrolases/*genetics/i;"
Notes:		"MedlineMizutani, Masaharu Nakanishi, Hidemitsu Ema, Jun-ichi Ma, Seung-Jin Noguchi, Etsuko Inohara-Ochiai, Misa Fukuchi-Mizutani, Masako Nakao, Masahiro Sakata, Kanzo eng Research Support, Non-U.S. Gov't 2002/12/14 Plant Physiol. 2002 Dec; 130(4):2164-76. doi: 10.1104/pp.102.011023"