Conformational Flexibility Enables the Function of a BECN1 Region Essential for Starvation-Mediated Autophagy

Biochemistry. 2016 Apr 5;55(13):1945-58. doi: 10.1021/acs.biochem.5b01264. Epub 2016 Mar 21.

Abstract

BECN1 is essential for autophagy, a critical eukaryotic cellular homeostasis pathway. Here we delineate a highly conserved BECN1 domain located between previously characterized BH3 and coiled-coil domains and elucidate its structure and role in autophagy. The 2.0 Å sulfur-single-wavelength anomalous dispersion X-ray crystal structure of this domain demonstrates that its N-terminal half is unstructured while its C-terminal half is helical; hence, we name it the flexible helical domain (FHD). Circular dichroism spectroscopy, double electron-electron resonance-electron paramagnetic resonance, and small-angle X-ray scattering (SAXS) analyses confirm that the FHD is partially disordered, even in the context of adjacent BECN1 domains. Molecular dynamic simulations fitted to SAXS data indicate that the FHD transiently samples more helical conformations. FHD helicity increases in 2,2,2-trifluoroethanol, suggesting it may become more helical upon binding. Lastly, cellular studies show that conserved FHD residues are required for starvation-induced autophagy. Thus, the FHD likely undergoes a binding-associated disorder-to-helix transition, and conserved residues critical for this interaction are essential for starvation-induced autophagy.

Publication types

  • 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 Sequence
  • Apoptosis Regulatory Proteins / chemistry*
  • Apoptosis Regulatory Proteins / genetics
  • Apoptosis Regulatory Proteins / metabolism
  • Autophagy*
  • Beclin-1
  • Biomarkers / metabolism
  • Cell Line, Tumor
  • Conserved Sequence
  • Culture Media, Serum-Free
  • Humans
  • Membrane Proteins / chemistry*
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Models, Molecular*
  • Molecular Docking Simulation
  • Molecular Dynamics Simulation
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism
  • Pliability
  • Point Mutation
  • Protein Conformation
  • Protein Refolding
  • Protein Stability
  • Protein Structure, Tertiary
  • Protein Unfolding
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism

Substances

  • Apoptosis Regulatory Proteins
  • BECN1 protein, human
  • Beclin-1
  • Biomarkers
  • Culture Media, Serum-Free
  • Membrane Proteins
  • Peptide Fragments
  • Recombinant Proteins