The ß-glucosidases responsible for bioactivation of hydroxynitrile glucosides in Lotus japonicus

Anne Vinther Morant; Nanna Bjarnholt; Mads Emil Kragh; Christian Hauge Kjærgaard; Kirsten Jørgensen; Suzanne Michelle Paquette; Markus Piotrowski; Anne Imberty; C Erik Olsen; Birger Lindberg Møller; Søren Bak

doi:10.1104/pp.107.109512

The ß-glucosidases responsible for bioactivation of hydroxynitrile glucosides in Lotus japonicus

Anne Vinther Morant, Nanna Bjarnholt, Mads Emil Kragh, Christian Hauge Kjærgaard, Kirsten Jørgensen, Suzanne Michelle Paquette, Markus Piotrowski, Anne Imberty, C Erik Olsen, Birger Lindberg Møller, Søren Bak

43 Citations (Scopus)

Abstract

Abstract: Lotus japonicus accumulates the hydroxynitrile glucosides lotaustralin, linamarin, and rhodiocyanosides A and D. Upon tissue disruption, the hydroxynitrile glucosides are bioactivated by hydrolysis by specific beta-glucosidases. A mixture of two hydroxynitrile glucoside-cleaving b-glucosidases was isolated from L. japonicus leaves and identified by protein sequencing as LjBGD2 and LjBGD4. The isolated hydroxynitrile glucoside-cleaving beta-glucosidases preferentially hydrolyzed rhodiocyanoside A and lotaustralin, whereas linamarin was only slowly hydrolyzed, in agreement with measurements of their rate of degradation upon tissue disruption in L. japonicus leaves. Comparative homology modeling predicted that LjBGD2 and LjBGD4 had nearly identical overall topologies and substrate-binding pockets. Heterologous expression of LjBGD2 and LjBGD4 in Arabidopsis (Arabidopsis thaliana) enabled analysis of their individual substrate specificity profiles and confirmed that both LjBGD2 and LjBGD4 preferentially hydrolyze the hydroxynitrile glucosides present in L. japonicus. Phylogenetic analyses revealed a third L. japonicus putative hydroxynitrile glucoside-cleaving beta-glucosidase, LjBGD7. Reverse transcription-polymerase chain reaction analysis showed that LjBGD2 and LjBGD4 are expressed in aerial parts of young L. japonicus plants, while LjBGD7 is expressed exclusively in roots. The differential expression pattern of LjBGD2, LjBGD4, and LjBGD7 corresponds to the previously observed expression profile for CYP79D3 and CYP79D4, encoding the two cytochromes P450 that catalyze the first committed step in the biosyntheis of hydroxynitrile glucosides in L. japonicus, with CYP79D3 expression in aerial tissues and CYP79D4 expression in roots.

Original language	English
Journal	Plant Physiology
Volume	147
Issue number	3
Pages (from-to)	1072-1091
Number of pages	20
ISSN	0032-0889
DOIs	https://doi.org/10.1104/pp.107.109512
Publication status	Published - 2008

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@article{27abda40c20211dd8ca2000ea68e967b,

title = "The {\ss}-glucosidases responsible for bioactivation of hydroxynitrile glucosides in Lotus japonicus",

abstract = "Abstract: Lotus japonicus accumulates the hydroxynitrile glucosides lotaustralin, linamarin, and rhodiocyanosides A and D. Upon tissue disruption, the hydroxynitrile glucosides are bioactivated by hydrolysis by specific beta-glucosidases. A mixture of two hydroxynitrile glucoside-cleaving b-glucosidases was isolated from L. japonicus leaves and identified by protein sequencing as LjBGD2 and LjBGD4. The isolated hydroxynitrile glucoside-cleaving beta-glucosidases preferentially hydrolyzed rhodiocyanoside A and lotaustralin, whereas linamarin was only slowly hydrolyzed, in agreement with measurements of their rate of degradation upon tissue disruption in L. japonicus leaves. Comparative homology modeling predicted that LjBGD2 and LjBGD4 had nearly identical overall topologies and substrate-binding pockets. Heterologous expression of LjBGD2 and LjBGD4 in Arabidopsis (Arabidopsis thaliana) enabled analysis of their individual substrate specificity profiles and confirmed that both LjBGD2 and LjBGD4 preferentially hydrolyze the hydroxynitrile glucosides present in L. japonicus. Phylogenetic analyses revealed a third L. japonicus putative hydroxynitrile glucoside-cleaving beta-glucosidase, LjBGD7. Reverse transcription-polymerase chain reaction analysis showed that LjBGD2 and LjBGD4 are expressed in aerial parts of young L. japonicus plants, while LjBGD7 is expressed exclusively in roots. The differential expression pattern of LjBGD2, LjBGD4, and LjBGD7 corresponds to the previously observed expression profile for CYP79D3 and CYP79D4, encoding the two cytochromes P450 that catalyze the first committed step in the biosyntheis of hydroxynitrile glucosides in L. japonicus, with CYP79D3 expression in aerial tissues and CYP79D4 expression in roots.",

author = "Morant, {Anne Vinther} and Nanna Bjarnholt and Kragh, {Mads Emil} and Kj{\ae}rgaard, {Christian Hauge} and Kirsten J{\o}rgensen and Paquette, {Suzanne Michelle} and Markus Piotrowski and Anne Imberty and Olsen, {C Erik} and M{\o}ller, {Birger Lindberg} and S{\o}ren Bak",

year = "2008",

doi = "10.1104/pp.107.109512",

language = "English",

volume = "147",

pages = "1072--1091",

journal = "Plant Physiology",

issn = "0032-0889",

publisher = "American Society of Plant Biologists",

number = "3",

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TY - JOUR

T1 - The ß-glucosidases responsible for bioactivation of hydroxynitrile glucosides in Lotus japonicus

AU - Morant, Anne Vinther

AU - Bjarnholt, Nanna

AU - Kragh, Mads Emil

AU - Kjærgaard, Christian Hauge

AU - Jørgensen, Kirsten

AU - Paquette, Suzanne Michelle

AU - Piotrowski, Markus

AU - Imberty, Anne

AU - Olsen, C Erik

AU - Møller, Birger Lindberg

AU - Bak, Søren

PY - 2008

Y1 - 2008

N2 - Abstract: Lotus japonicus accumulates the hydroxynitrile glucosides lotaustralin, linamarin, and rhodiocyanosides A and D. Upon tissue disruption, the hydroxynitrile glucosides are bioactivated by hydrolysis by specific beta-glucosidases. A mixture of two hydroxynitrile glucoside-cleaving b-glucosidases was isolated from L. japonicus leaves and identified by protein sequencing as LjBGD2 and LjBGD4. The isolated hydroxynitrile glucoside-cleaving beta-glucosidases preferentially hydrolyzed rhodiocyanoside A and lotaustralin, whereas linamarin was only slowly hydrolyzed, in agreement with measurements of their rate of degradation upon tissue disruption in L. japonicus leaves. Comparative homology modeling predicted that LjBGD2 and LjBGD4 had nearly identical overall topologies and substrate-binding pockets. Heterologous expression of LjBGD2 and LjBGD4 in Arabidopsis (Arabidopsis thaliana) enabled analysis of their individual substrate specificity profiles and confirmed that both LjBGD2 and LjBGD4 preferentially hydrolyze the hydroxynitrile glucosides present in L. japonicus. Phylogenetic analyses revealed a third L. japonicus putative hydroxynitrile glucoside-cleaving beta-glucosidase, LjBGD7. Reverse transcription-polymerase chain reaction analysis showed that LjBGD2 and LjBGD4 are expressed in aerial parts of young L. japonicus plants, while LjBGD7 is expressed exclusively in roots. The differential expression pattern of LjBGD2, LjBGD4, and LjBGD7 corresponds to the previously observed expression profile for CYP79D3 and CYP79D4, encoding the two cytochromes P450 that catalyze the first committed step in the biosyntheis of hydroxynitrile glucosides in L. japonicus, with CYP79D3 expression in aerial tissues and CYP79D4 expression in roots.

AB - Abstract: Lotus japonicus accumulates the hydroxynitrile glucosides lotaustralin, linamarin, and rhodiocyanosides A and D. Upon tissue disruption, the hydroxynitrile glucosides are bioactivated by hydrolysis by specific beta-glucosidases. A mixture of two hydroxynitrile glucoside-cleaving b-glucosidases was isolated from L. japonicus leaves and identified by protein sequencing as LjBGD2 and LjBGD4. The isolated hydroxynitrile glucoside-cleaving beta-glucosidases preferentially hydrolyzed rhodiocyanoside A and lotaustralin, whereas linamarin was only slowly hydrolyzed, in agreement with measurements of their rate of degradation upon tissue disruption in L. japonicus leaves. Comparative homology modeling predicted that LjBGD2 and LjBGD4 had nearly identical overall topologies and substrate-binding pockets. Heterologous expression of LjBGD2 and LjBGD4 in Arabidopsis (Arabidopsis thaliana) enabled analysis of their individual substrate specificity profiles and confirmed that both LjBGD2 and LjBGD4 preferentially hydrolyze the hydroxynitrile glucosides present in L. japonicus. Phylogenetic analyses revealed a third L. japonicus putative hydroxynitrile glucoside-cleaving beta-glucosidase, LjBGD7. Reverse transcription-polymerase chain reaction analysis showed that LjBGD2 and LjBGD4 are expressed in aerial parts of young L. japonicus plants, while LjBGD7 is expressed exclusively in roots. The differential expression pattern of LjBGD2, LjBGD4, and LjBGD7 corresponds to the previously observed expression profile for CYP79D3 and CYP79D4, encoding the two cytochromes P450 that catalyze the first committed step in the biosyntheis of hydroxynitrile glucosides in L. japonicus, with CYP79D3 expression in aerial tissues and CYP79D4 expression in roots.

U2 - 10.1104/pp.107.109512

DO - 10.1104/pp.107.109512

M3 - Journal article

C2 - 18467457

SN - 0032-0889

VL - 147

SP - 1072

EP - 1091

JO - Plant Physiology

JF - Plant Physiology

IS - 3

ER -

The ß-glucosidases responsible for bioactivation of hydroxynitrile glucosides in Lotus japonicus

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