Studies of biomolecular conformations and conformational dynamics by mass spectrometry

 I. Introduction 38A. Protein Conformations and Dynamics 38B. 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 40C. Folding and Binding 41 II. Mass Spectrometry‐Based Approaches to Study Protein Conformations and Dynamics 41A. Positive‐Ion Charge‐State Distributions and Biomolecular Shapes in Solution (ESI MS) 41B. Amide Hydrogen Exchange 44   1. Mechanism 44   2. HDX Pulse Labeling 45   3. Site‐Specific Measurement of HDX by Protein Fragmentation 46C. Differential Chemical Labeling 49D. Gas‐Phase Methods of Probing Multi‐Protein Complex Topology 53 III. Protein‐Folding Studies 53A. Characterization of Kinetic Intermediates in Refolding Experiments 53B. Characterization of Equilibrium Intermediates in Unfolding Experiments 55 IV. Studies of Protein‐Structure and Dynamics Under Native Conditions 58A. Characterization of Higher‐Order Structure 58   1. Quaternary Structure 58   2. Secondary and Tertiary Structure 60B. Probing Structural Dynamics Under Native Conditions 60C. Ligand‐ and Substrate‐Induced Conformational Changes in Proteins 62V. Conclusions 65A. Place of MS in Experimental Biophysics: Uniqueness and Complementarity to Other Techniques 65B. Future Directions in the Field: Perspectives and Challenges 65 Acknowledgments 66 References 66In 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