Time-resolved neutron scattering provides new insight into protein substrate processing by a AAA+ unfoldase

Sci Rep. 2017 Jan 19;7:40948. doi: 10.1038/srep40948.


We present a combination of small-angle neutron scattering, deuterium labelling and contrast variation, temperature activation and fluorescence spectroscopy as a novel approach to obtain time-resolved, structural data individually from macromolecular complexes and their substrates during active biochemical reactions. The approach allowed us to monitor the mechanical unfolding of a green fluorescent protein model substrate by the archaeal AAA+ PAN unfoldase on the sub-minute time scale. Concomitant with the unfolding of its substrate, the PAN complex underwent an energy-dependent transition from a relaxed to a contracted conformation, followed by a slower expansion to its initial state at the end of the reaction. The results support a model in which AAA ATPases unfold their substrates in a reversible power stroke mechanism involving several subunits and demonstrate the general utility of this time-resolved approach for studying the structural molecular kinetics of multiple protein remodelling complexes and their substrates on the sub-minute time scale.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATPases Associated with Diverse Cellular Activities / chemistry*
  • ATPases Associated with Diverse Cellular Activities / metabolism*
  • Archaea / enzymology*
  • Green Fluorescent Proteins / metabolism
  • Protein Conformation
  • Protein Folding
  • Scattering, Small Angle*


  • Green Fluorescent Proteins
  • ATPases Associated with Diverse Cellular Activities