Carbohydrate-Binding Capability and Functional Conformational Changes of AbnE, an Arabino-oligosaccharide Binding Protein

Shifra Lansky, Rachel Salama, Smadar Shulami, Noa Lavid, Saumik Sen, Igor Schapiro, Yuval Shoham, Gil Shoham*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

ABC importers are membrane proteins responsible for the transport of nutrients into the cells of prokaryotes. Although the structures of ABC importers vary, all contain four conserved domains: two nucleotide-binding domains (NBDs), which bind and hydrolyze ATP, and two transmembrane domains (TMDs), which help translocate the substrate. ABC importers are also dependent on an additional protein component, a high-affinity substrate-binding protein (SBP) that specifically binds the target ligand for delivery to the appropriate ABC transporter. AbnE is a SBP belonging to the ABC importer for arabino-oligosaccharides in the Gram-positive thermophilic bacterium Geobacillus stearothermophilus. Using isothermal titration calorimetry (ITC), purified AbnE was shown to bind medium-sized arabino-oligosaccharides, in the range of arabino-triose (A3) to arabino-octaose (A8), all with Kd values in the nanomolar range. We describe herein the 3D structure of AbnE in its closed conformation in complex with a wide range of arabino-oligosaccharide substrates (A2-A8). These structures provide the basis for the detailed structural analysis of the AbnE-sugar complexes, and together with complementary quantum chemical calculations, site-specific mutagenesis, and isothermal titration calorimetry (ITC) experiments, provide detailed insights into the AbnE-substrate interactions involved. Small-angle X-ray scattering (SAXS) experiments and normal mode analysis (NMA) are used to study the conformational changes of AbnE, and these data, taken together, suggest clues regarding its binding mode to the full ABC importer.

Original languageAmerican English
Pages (from-to)2099-2120
Number of pages22
JournalJournal of Molecular Biology
Volume432
Issue number7
DOIs
StatePublished - 27 Mar 2020

Bibliographical note

Publisher Copyright:
© 2020

Keywords

  • ABC transporter
  • Arabinan utilization system
  • Geobacillus stearothermophilus
  • arabinose
  • substrate-binding protein

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