Plant α-glucan phosphatases SEX4 and LSF2 display different affinity for amylopectin and amylose

FEBS Lett. 2016 Jan;590(1):118-28. doi: 10.1002/1873-3468.12027. Epub 2016 Jan 4.

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.

Keywords: Like Sex Four2; Starch Excess 4; affinity gel electrophoresis; carbohydrate binding domain; surface binding sites; surface plasmon resonance.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Substitution
  • Amylopectin / chemistry
  • Amylopectin / metabolism*
  • Amylose / chemistry
  • Amylose / metabolism*
  • Arabidopsis / enzymology*
  • Arabidopsis Proteins / chemistry
  • Arabidopsis Proteins / genetics
  • Arabidopsis Proteins / metabolism*
  • Binding Sites
  • Carbohydrate Conformation
  • Cytoplasmic Granules / chemistry
  • Cytoplasmic Granules / enzymology
  • Cytoplasmic Granules / metabolism
  • Dual-Specificity Phosphatases / chemistry
  • Dual-Specificity Phosphatases / genetics
  • Dual-Specificity Phosphatases / metabolism*
  • Kinetics
  • Models, Molecular*
  • Molecular Dynamics Simulation
  • Mutation
  • Plant Leaves / enzymology*
  • Protein Conformation
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Substrate Specificity
  • Surface Plasmon Resonance
  • beta-Cyclodextrins / chemistry
  • beta-Cyclodextrins / metabolism

Substances

  • Arabidopsis Proteins
  • Recombinant Proteins
  • beta-Cyclodextrins
  • Amylose
  • Amylopectin
  • Dual-Specificity Phosphatases
  • LSF2 protein, Arabidopsis
  • SEX4 protein, Arabidopsis
  • betadex

Associated data

  • GENBANK/AT3G10940
  • GENBANK/AT3G52180
  • GENBANK/M73321