In vitro characterization of the homogalacturonan binding domain of the wall-associated kinase WAK1 using site-directed mutagenesis

Annabelle Decreux, Annick THOMAS, Benoît SPIES, Robert BRASSEUR, Pierre Van Cutsem, Johan Messiaen

Research output: Contribution to journalArticle

Abstract

Wall-associated kinase 1 ' WAK1 is a transmembrane protein containing a cytoplasmic Ser/Thr kinase domain and an extracellular domain in contact with the pectin fraction of the plant cell wall in Arabidopsis thaliana (L.) Heynh. In a previous paper (Decreux and Messiaen, 2005), we showed that a recombinant peptide expressed in yeast corresponding to amino acids 67 to 254 of the extracellular domain of WAK1 specifically interacts with commercial non-methylesterified homogalacturonic acid, purified homogalacturonans from Arabidopsis and oligogalacturonides in a calcium-induced conformation. In this report, we used a receptor binding domain sequence-based prediction method to identify four putative binding sites in the extracellular domain of WAK1, in which cationic amino acids were selected for substitution by site-directed mutagenesis. Interaction studies between mutated forms of WAK1 and homogalacturonans allowed us to identify and confirm at least five specific amino acids involved in the interaction with homogalacturonan dimers and multimers. The presence of this homogalacturonan binding domain within the extracellular domain of WAK1 is discussed in terms of cell wall architecture and signal transduction.
Original languageEnglish
Pages (from-to)1068-1079
Number of pages12
JournalPhytochemistry
Volume67
Issue number11
Publication statusPublished - 2006

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Mutagenesis
site-directed mutagenesis
Site-Directed Mutagenesis
phosphotransferases (kinases)
Phosphotransferases
Arabidopsis
Amino Acids
Cell Wall
amino acids
cell walls
Signal transduction
transmembrane proteins
Plant Cells
Dimers
Yeast
pectins
signal transduction
Conformations
binding sites
Signal Transduction

Cite this

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title = "In vitro characterization of the homogalacturonan binding domain of the wall-associated kinase WAK1 using site-directed mutagenesis",
abstract = "Wall-associated kinase 1 ' WAK1 is a transmembrane protein containing a cytoplasmic Ser/Thr kinase domain and an extracellular domain in contact with the pectin fraction of the plant cell wall in Arabidopsis thaliana (L.) Heynh. In a previous paper (Decreux and Messiaen, 2005), we showed that a recombinant peptide expressed in yeast corresponding to amino acids 67 to 254 of the extracellular domain of WAK1 specifically interacts with commercial non-methylesterified homogalacturonic acid, purified homogalacturonans from Arabidopsis and oligogalacturonides in a calcium-induced conformation. In this report, we used a receptor binding domain sequence-based prediction method to identify four putative binding sites in the extracellular domain of WAK1, in which cationic amino acids were selected for substitution by site-directed mutagenesis. Interaction studies between mutated forms of WAK1 and homogalacturonans allowed us to identify and confirm at least five specific amino acids involved in the interaction with homogalacturonan dimers and multimers. The presence of this homogalacturonan binding domain within the extracellular domain of WAK1 is discussed in terms of cell wall architecture and signal transduction.",
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journal = "Phytochemistry",
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In vitro characterization of the homogalacturonan binding domain of the wall-associated kinase WAK1 using site-directed mutagenesis. / Decreux, Annabelle; THOMAS, Annick; SPIES, Benoît; BRASSEUR, Robert; Van Cutsem, Pierre; Messiaen, Johan.

In: Phytochemistry , Vol. 67, No. 11, 2006, p. 1068-1079.

Research output: Contribution to journalArticle

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T1 - In vitro characterization of the homogalacturonan binding domain of the wall-associated kinase WAK1 using site-directed mutagenesis

AU - Decreux, Annabelle

AU - THOMAS, Annick

AU - SPIES, Benoît

AU - BRASSEUR, Robert

AU - Van Cutsem, Pierre

AU - Messiaen, Johan

PY - 2006

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AB - Wall-associated kinase 1 ' WAK1 is a transmembrane protein containing a cytoplasmic Ser/Thr kinase domain and an extracellular domain in contact with the pectin fraction of the plant cell wall in Arabidopsis thaliana (L.) Heynh. In a previous paper (Decreux and Messiaen, 2005), we showed that a recombinant peptide expressed in yeast corresponding to amino acids 67 to 254 of the extracellular domain of WAK1 specifically interacts with commercial non-methylesterified homogalacturonic acid, purified homogalacturonans from Arabidopsis and oligogalacturonides in a calcium-induced conformation. In this report, we used a receptor binding domain sequence-based prediction method to identify four putative binding sites in the extracellular domain of WAK1, in which cationic amino acids were selected for substitution by site-directed mutagenesis. Interaction studies between mutated forms of WAK1 and homogalacturonans allowed us to identify and confirm at least five specific amino acids involved in the interaction with homogalacturonan dimers and multimers. The presence of this homogalacturonan binding domain within the extracellular domain of WAK1 is discussed in terms of cell wall architecture and signal transduction.

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