期刊
PROTEIN SCIENCE
卷 24, 期 8, 页码 1301-1312出版社
WILEY
DOI: 10.1002/pro.2706
关键词
ion mobility mass spectrometry; protein dynamics; drug discovery; 1-antitrypsin; protein unfolding; mass spectrometry; methods; nuclear magnetic resonance; biomolecular; X-ray crystallography
资金
- Wellcome Trust
- Alpha-1 Foundation
- A MRC PhD studentship [MR/J003867/1]
- MRC
- UCLH NIHR Biomedical Research Centre
- GlaxoSmithKline
- Wellcome Trust Intermediate Fellowship
- The Alpha-1 Foundation
- Medical Research Council [G0901786] Funding Source: researchfish
- National Institute for Health Research [NF-SI-0513-10070] Funding Source: researchfish
- MRC [G0901786] Funding Source: UKRI
Native mass spectrometry (MS) methods permit the study of multiple protein species within solution equilibria, whereas ion mobility (IM)-MS can report on conformational behavior of specific states. We used IM-MS to study a conformationally labile protein ((1)-antitrypsin) that undergoes pathological polymerization in the context of point mutations. The folded, native state of the Z-variant remains highly polymerogenic in physiological conditions despite only minor thermodynamic destabilization relative to the wild-type variant. Various data implicate kinetic instability (conformational lability within a native state ensemble) as the basis of Z (1)-antitrypsin polymerogenicity. We show the ability of IM-MS to track such disease-relevant conformational behavior in detail by studying the effects of peptide binding on (1)-antitrypsin conformation and dynamics. IM-MS is, therefore, an ideal platform for the screening of compounds that result in therapeutically beneficial kinetic stabilization of native (1)-antitrypsin. Our findings are confirmed with high-resolution X-ray crystallographic and nuclear magnetic resonance spectroscopic studies of the same event, which together dissect structural changes from dynamic effects caused by peptide binding at a residue-specific level. IM-MS methods, therefore, have great potential for further study of biologically relevant thermodynamic and kinetic instability of proteins and provide rapid and multidimensional characterization of ligand interactions of therapeutic interest.
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