title: Plant α-glucan phosphatases SEX4 and LSF2 display different affinity for amylopectin and amylose.
abstract: The plant glucan phosphatases Starch EXcess 4 (SEX4) and Like Sex Four2 (LSF2) apply different starch binding mechanisms. SEX4 contains a carbohydrate binding module, and LSF2 has two surface binding sites (SBSs). We determined KDapp for amylopectin and amylose, and KD for β-cyclodextrin and validated binding site mutants deploying affinity gel electrophoresis (AGE) and surface plasmon resonance. SEX4 has a higher affinity for amylopectin; LSF2 prefers amylose and β-cyclodextrin. SEX4 has 50-fold lower KDapp for amylopectin compared to LSF2. Molecular dynamics simulations and AGE data both support long-distance mutual effects of binding at SBSs and the active site in LSF2.
authors.affiliation: Enzyme and Protein Chemistry, Department of Systems Biology, Technical University of Denmark, Kongens Lyngby, Denmark.
authors.forename: Casper
authors.initials: C
authors.lastname: Wilkens
authors.affiliation: Department of Molecular and Cellular Biochemistry and Center for Structural Biology, University of Kentucky, Lexington, KY, USA.
authors.forename: Kyle D
authors.initials: KD
authors.lastname: Auger
authors.affiliation: Department of Chemical and Materials Engineering, University of Kentucky, Lexington, KY, USA.
authors.forename: Nolan T
authors.initials: NT
authors.lastname: Anderson
authors.affiliation: Department of Molecular and Cellular Biochemistry and Center for Structural Biology, University of Kentucky, Lexington, KY, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Meekins
authors.affiliation: Department of Molecular and Cellular Biochemistry and Center for Structural Biology, University of Kentucky, Lexington, KY, USA.
authors.forename: Madushi
authors.initials: M
authors.lastname: Raththagala
authors.affiliation: Enzyme and Protein Chemistry, Department of Systems Biology, Technical University of Denmark, Kongens Lyngby, Denmark.
authors.forename: Maher
authors.initials: M
authors.lastname: Abou Hachem
authors.affiliation: Department of Chemical and Materials Engineering, University of Kentucky, Lexington, KY, USA.
authors.forename: Christina M
authors.initials: CM
authors.lastname: Payne
authors.affiliation: Department of Molecular and Cellular Biochemistry and Center for Structural Biology, University of Kentucky, Lexington, KY, USA.
authors.forename: Matthew S
authors.initials: MS
authors.lastname: Gentry
authors.affiliation: Enzyme and Protein Chemistry, Department of Systems Biology, Technical University of Denmark, Kongens Lyngby, Denmark.
authors.forename: Birte
authors.initials: B
authors.lastname: Svensson
mesh_terms: D019943:Amino Acid Substitution; D000687:Amylopectin / Q000737:chemistry / Q000378:metabolism*; D000688:Amylose / Q000737:chemistry / Q000378:metabolism*; D017360:Arabidopsis / Q000201:enzymology*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002236:Carbohydrate Conformation; D003594:Cytoplasmic Granules / Q000737:chemistry / Q000201:enzymology / Q000378:metabolism; D054637:Dual-Specificity Phosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D009154:Mutation; D018515:Plant Leaves / Q000201:enzymology*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D020349:Surface Plasmon Resonance; D047392:beta-Cyclodextrins / Q000737:chemistry / Q000378:metabolism
chemical_list: D000687:Amylopectin; D000688:Amylose; D029681:Arabidopsis Proteins; D054637:Dual-Specificity Phosphatases; D011994:Recombinant Proteins; D047392:beta-Cyclodextrins; C574400:LSF2 protein, Arabidopsis; C553186:At3g52180 protein, Arabidopsis; C031215:betadex
keywords: Like Sex Four2; Starch Excess 4; affinity gel electrophoresis; carbohydrate binding domain; surface binding sites; surface plasmon resonance