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

Casper Wilkens, Kyle D. Auger, Nolan T. Anderson, David A. Meekins, Madushi Raththagala, Maher Abou Hachem, Christina M. Payne, Matthew S. Gentry, Birte Svensson

Research output: Contribution to journalArticlepeer-review

15 Scopus citations

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.

Original languageEnglish
Pages (from-to)118-128
Number of pages11
JournalFEBS Letters
Volume590
Issue number1
DOIs
StatePublished - Jan 2016

Bibliographical note

Funding Information:
This work was supported by The Danish Council for Independent Research | Natural Sciences (FNU) to the project ‘Discovery and Characterization of Carbohydrate Surface Binding Sites (SBS) in Polysaccharide Converting Enzymes’ and the Biacore T100 instrument (both to BS) and a National Science Foundation CAREER Grant MCB-1252345, National Institutes of Health Grant P20GM103486, Kentucky Science and Energy Foundation Grant KSE-2971-RDE-0147, Mizutani Foundation for Glycoscience Award, and NSF Grant IIA-1355438 (all to MSG). The Technical University of Denmark co-financed a PhD fellowship (CW). Computational resources were provided by the University of Kentucky and the Extreme Science and Engineering Discovery Environment (XSEDE) supported by National Science Foundation grant number ACI-1053575 (allocation TG-MCB090159). Andreas Blennow, University of Copenhagen is thanked for fruitful discussions on the structure of phosphorylated starch and David Wilson, Cornell University for the T. fusca Cel6A.

Publisher Copyright:
© 2015 Federation of European Biochemical Societies.

Keywords

  • Like Sex Four2
  • Starch Excess 4
  • affinity gel electrophoresis
  • carbohydrate binding domain
  • surface binding sites
  • surface plasmon resonance

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

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