posted on 2011-10-11, 00:00authored byTheodore
R. Keppel, Brent A. Howard, David D. Weis
Mapping the structured and disordered regions and identifying
disorder-to-order
transitions are essential to understanding intrinsically disordered
proteins (IDPs). One technique that can provide such information is
H/D exchange coupled with mass spectrometry (H/D-MS). To explore the
feasibility of H/D-MS for mapping disordered and ordered regions in
IDPs, we undertook a systematic evaluation of an unstructured protein,
a molten globular protein, and the well-folded complex of the two
proteins. Most segments of the unstructured protein, ACTR (activator
of thyroid and retinoid receptors, NCOA3_HUMAN, residues 1018–1088),
exchange at rates consistent with its assignment as an unstructured
protein, but there is slight protection in regions that become helical
in the ACTR–CBP complex. The molten globular protein, CBP (the
nuclear coactivator binding domain of the CREB binding protein, CBP_MOUSE,
residues 2059–2117), is moderately protected from exchange,
and the protection is nearly uniform across the length of the protein.
The uniformity arises because of rapid interconversion between an
ensemble of folded conformers and an ensemble of unstructured conformers.
Rapid interconversion causes the H/D exchange kinetics to be dominated
by exchange by molecules in unstructured conformations. For the folded
ACTR–CBP complex, the exchange data provide a qualitatively
accurate description of the complex. Our results provide a useful
framework to use in the interpretation of H/D-MS data of intrinsically
disordered proteins.