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Updated: May 31, 2026
Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics
Published on: April 17, 2017
Complexation and Calcium-Induced Conformational Changes in the Cardiac Troponin Complex Monitored by
George M Bou-Assaf1, Jean E Chamoun, Mark R Emmett
1Department of Chemistry and Biochemistry, Florida State University, Tallahassee, FL 32306.
Insights
Cardiac troponin
Area of Science:
- Biochemistry
- Structural Biology
- Molecular Cardiology
Background:
- Cardiac muscle contraction relies on the troponin complex.
- Understanding troponin's structure and dynamics is key to cardiac function.
Purpose of the Study:
- To investigate the structural dynamics of cardiac troponin.
- To map Ca(2+)-induced conformational changes within the troponin complex.
Main Methods:
- Solution-phase hydrogen/deuterium exchange (HDX).
- Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR MS).
- Comparative analysis of troponin C (TnC) alone, TnC:TnI, and TnC:TnI:TnT complexes.
Main Results:
- Identified flexible and rigid domains within the troponin complex.
- Observed fast exchange rates in TnI's N-terminal extension and TnC's DE linker.
- Slowest exchange rates in the stable IT coiled-coil domain.
- Ca(2+) binding to TnC's N-lobe induced allosteric effects, protecting the C-lobe and transmitting changes to the IT coiled-coil and TnT1.
Conclusions:
- Cardiac troponin exhibits distinct flexible and rigid regions.
- Calcium binding triggers long-range conformational changes throughout the complex.
- HDX-MS provides insights into troponin dynamics, complementing existing structural data.
Abstract:
Cardiac muscle contraction is regulated by the heterotrimeric complex: troponin. We apply solution-phase hydrogen/deuterium exchange monitored by FT-ICR mass spectrometry to study the structural dynamics and the Ca-induced conformational changes of the cardiac isoform of troponin, by comparing H/D exchange rate constants for TnC alone, the binary TnC:TnI complex, and the ternary TnC:TnI:TnT complex for Ca-free and Ca-saturated states. The wide range of exchange rate constants indicates that the complexes possess both highly flexible and very rigid domains. Fast exchange rates were observed for the N-terminal extension of TnI (specific to the cardiac isoform), the DE linker in TnC alone, and the mobile domain of TnI. The slowest rates were for the IT coiled-coil that grants stability and stiffness to the complex. Ca(2+) binding to site II of the N-lobe of TnC induces short-range allosteric effects, mainly protection for the C-lobe of TnC that transmits long-range conformational changes that reach the IT coiled-coil and even TnT1. The present results corroborate prior X-ray crystallography and NMR interpretations and also illuminate domains that were not resolved or truncated in those experiments.
