Studies of biomolecular conformations and conformational dynamics by mass spectrometry
Authors: Kaltashov Igor A. , Eyles Stephen J. 
Published: October 2002

Abstract

I.
Introduction
38

A. Protein Conformations and Dynamics
38

B. Protein‐Energy Landscapes: The Folding Problem and Dynamics in the Native State
38

1. Protein Conformational Ensembles and Energy Landscapes: Enthalpic and Entropic Considerations
38

2. Kinetic and Equilibrium Intermediates
39

3. Definition and Specific Examples of Protein Conformations: Native Conformations, Random Coils, Late‐Folding Intermediates, and Molten Globules
40

C. Folding and Binding
41

II.
Mass Spectrometry‐Based Approaches to Study Protein Conformations and Dynamics
41

A. Positive‐Ion Charge‐State Distributions and Biomolecular Shapes in Solution (ESI MS)
41

B. Amide Hydrogen Exchange
44

1. Mechanism
44

2. HDX Pulse Labeling
45

3. Site‐Specific Measurement of HDX by Protein Fragmentation
46

C. Differential Chemical Labeling
49

D. Gas‐Phase Methods of Probing Multi‐Protein Complex Topology
53

III.
Protein‐Folding Studies
53

A. Characterization of Kinetic Intermediates in Refolding Experiments
53

B. Characterization of Equilibrium Intermediates in Unfolding Experiments
55

IV.
Studies of Protein‐Structure and Dynamics Under Native Conditions
58

A. Characterization of Higher‐Order Structure
58

1. Quaternary Structure
58

2. Secondary and Tertiary Structure
60

B. Probing Structural Dynamics Under Native Conditions
60

C. Ligand‐ and Substrate‐Induced Conformational Changes in Proteins
62

V. Conclusions
65

A. Place of MS in Experimental Biophysics: Uniqueness and Complementarity to Other Techniques
65

B. Future Directions in the Field: Perspectives and Challenges
65

Acknowledgments
66

References
66

In the post‐genomic era, a wealth of structural information has been amassed for proteins from NMR and crystallography. However, static protein structures alone are not a sufficient description: knowledge of the dynamic nature of proteins is essential to understand their wide range of functions and behavior during the life cycle from synthesis to degradation. Furthermore, few proteins have the ability to act alone in the crowded cellular environment. Assemblies of multiple proteins governed by complex signaling pathways are often required for the tasks of target recognition, binding, transport, and function. Mass spectrometry has emerged over the past several years as a powerful tool to address many of these questions. Recent improvements in “soft” ionization techniques have enabled researchers to study proteins and biomolecular complexes, both directly and indirectly. Likewise, continuous improvements in instrumental design in recent years have resulted in a dramatic expansion of the m/z range and resolution, enabling observation of large multi‐protein assemblies whose structures are retained in the gas phase. In this article, we discuss some of the mass spectrometric techniques applied to investigate the nature of the conformations and dynamical properties that govern protein function. © 2002 Wiley Periodicals, Inc. Mass Spec Rev 21:37–71, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/mas.10017