title: The crystal structure of the thiocyanate-forming protein from Thlaspi arvense, a kelch protein involved in glucosinolate breakdown.
abstract: Kelch repeat-containing proteins are involved in diverse cellular processes, but only a small subset of plant kelch proteins has been functionally characterized. Thiocyanate-forming protein (TFP) from field-penny cress, Thlaspi arvense (Brassicaceae), is a representative of specifier proteins, a group of kelch proteins involved in plant specialized metabolism. As components of the glucosinolate-myrosinase system of the Brassicaceae, specifier proteins determine the profile of bioactive products formed when plant tissue is disrupted and glucosinolates are hydrolyzed by myrosinases. Here, we describe the crystal structure of TaTFP at a resolution of 1.4 Å. TaTFP crystallized as homodimer. Each monomer forms a six-blade β-propeller with a wide "top" and a narrower "bottom" opening with distinct strand-connecting loops protruding far beyond the lower propeller surface. Molecular modeling and mutational analysis identified residues for glucosinolate aglucone and Fe(2+) cofactor binding within these loops. As the first experimentally determined structure of a plant kelch protein, the crystal structure of TaTFP not only enables more detailed mechanistic studies on glucosinolate breakdown product formation, but also provides a new basis for research on the diverse roles and mechanisms of other kelch proteins in plants.
authors.affiliation: Institute of Pharmaceutical Biology, Technische Universität Braunschweig, Mendelssohnstr. 1, 38106, Braunschweig, Germany.
authors.forename: Frauke
authors.initials: F
authors.lastname: Gumz
authors.affiliation: Structure and Function of Proteins, Helmholtz Center for Infection Research, Inhoffenstr. 7, 38124, Braunschweig, Germany.
authors.forename: Joern
authors.initials: J
authors.lastname: Krausze
authors.affiliation: Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, 06120, Halle (Saale), Germany.
authors.forename: Daniela
authors.initials: D
authors.lastname: Eisenschmidt
authors.affiliation: Institute of Pharmaceutical Biology, Technische Universität Braunschweig, Mendelssohnstr. 1, 38106, Braunschweig, Germany.
authors.forename: Anita
authors.initials: A
authors.lastname: Backenköhler
authors.affiliation: Institute of Pharmaceutical Biology, Technische Universität Braunschweig, Mendelssohnstr. 1, 38106, Braunschweig, Germany.
authors.forename: Leif
authors.initials: L
authors.lastname: Barleben
authors.affiliation: Department of Bioorganic Chemistry, Leibniz Institute of Plant Biochemistry, Weinberg 3, 06120, Halle (Saale), Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Brandt
authors.affiliation: Institute of Pharmaceutical Biology, Technische Universität Braunschweig, Mendelssohnstr. 1, 38106, Braunschweig, Germany. u.wittstock@tu-bs.de.
authors.forename: Ute
authors.initials: U
authors.lastname: Wittstock
mesh_terms: D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D005961:Glucosinolates / Q000378:metabolism*; D062105:Molecular Docking Simulation; D010940:Plant Proteins / Q000737:chemistry* / Q000502:physiology; D017434:Protein Structure, Tertiary; D013861:Thiocyanates / Q000378:metabolism; D031227:Thlaspi / Q000378:metabolism / Q000502:physiology*
chemical_list: D005961:Glucosinolates; D010940:Plant Proteins; D013861:Thiocyanates; C031760:thiocyanate
keywords: Glucosinolate breakdown; Glucosinolate hydrolysis; Kelch motif; Molecular modeling; Site-directed mutagenesis; Specifier protein