Mapping residual structure in intrinsically disordered proteins at residue resolution using millisecond hydrogen/deuterium exchange and residue averaging

J Am Soc Mass Spectrom. 2015 Apr;26(4):547-54. doi: 10.1007/s13361-014-1033-6. Epub 2014 Dec 7.

Abstract

Measurement of residual structure in intrinsically disordered proteins can provide insights into the mechanisms by which such proteins undergo coupled binding and folding. The present work describes an approach to measure residual structure in disordered proteins using millisecond hydrogen/deuterium (H/D) exchange in a conventional bottom-up peptide-based workflow. We used the exchange mid-point, relative to a totally deuterated control, to quantify the rate of H/D exchange in each peptide. A weighted residue-by-residue average of these midpoints was used to map the extent of residual structure at near single-residue resolution. We validated this approach both by simulating a disordered protein and experimentally using the p300 binding domain of ACTR, a model disordered protein already well-characterized by other approaches. Secondary structure elements mapped in the present work are in good agreement with prior nuclear magnetic resonance measurements. The new approach was somewhat limited by a loss of spatial resolution and subject to artifacts because of heterogeneities in intrinsic exchange. Approaches to correct these limitations are discussed.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Deuterium Exchange Measurement / methods*
  • Intrinsically Disordered Proteins / analysis*
  • Intrinsically Disordered Proteins / chemistry*
  • Mass Spectrometry / methods*
  • Peptide Mapping / methods*
  • Protein Structure, Secondary

Substances

  • Intrinsically Disordered Proteins