title: Cassia obtusifolia MetE as a cytosolic target for potassium isolespedezate, a leaf-opening factor of Cassia plants: target exploration by a compact molecular-probe strategy.
abstract: Affinity chromatography by using ligand-immobilized bead technology is generally the first choice for target exploration of a bioactive ligand. However, when a ligand has comparatively low affinity against its target, serious difficulties will be raised in affinity-based target detection. We report here that the use of compact molecular probes (CMP) will be advantageous in such cases; it enables the retention of moderate affinity between the ligand and its target in contrast to immobilizing the ligand on affinity beads that will cause a serious drop in affinity to preclude target detection. In the CMP strategy, a CMP containing an azide handle is used for an initial affinity-based labeling of target, and subsequent tagging by CuAAC with a large FLAG tag will give a tagged target protein. By using the CMP strategy, we succeeded in the identification of Cassia obtusifolia MetE as a cytosolic target protein of potassium isolespedezate (1), a moderately bioactive ligand.
authors.affiliation: Department of Chemistry, Tohoku University, 6-3 Aramaki-aza Aoba, Sendai 980-8578, Japan. ueda@m.tohoku.ac.jp
authors.forename: Minoru
authors.initials: M
authors.lastname: Ueda
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Manabe
authors.forename: Yuki
authors.initials: Y
authors.lastname: Otsuka
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Kanzawa
mesh_terms: D000595:Amino Acid Sequence; D002366:Cassia / Q000201:enzymology*; D002846:Chromatography, Affinity; D003373:Coumaric Acids / Q000737:chemistry / Q000378:metabolism*; D005960:Glucosides / Q000737:chemistry / Q000378:metabolism*; D008780:Methyltransferases / Q000032:analysis / Q000378:metabolism*; D015335:Molecular Probes / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000502:physiology; D010940:Plant Proteins / Q000032:analysis / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: D003373:Coumaric Acids; D005960:Glucosides; D015335:Molecular Probes; D010940:Plant Proteins; C558667:isolespedezic acid; D008780:Methyltransferases