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T-wave Ion Mobility-mass Spectrometry: Basic Experimental Procedures for Protein Complex Analysis
Published on: July 31, 2010
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Following Structural Changes by Thermal Denaturation Using Trapped Ion Mobility Spectrometry-Mass Spectrometry
Kevin Jeanne Dit Fouque1, Francisco Fernandez-Lima1,2
1Department of Chemistry and Biochemistry, Florida International University, Miami, Florida 33199, United States.
The Journal of Physical Chemistry. B
|June 21, 2020
Summary
This study used trapped ion mobility spectrometry-mass spectrometry to analyze bovine serum albumin (BSA) structural changes with temperature. It revealed multiple transitions beyond a simple two-state unfolding, offering a detailed view of protein behavior.
Area of Science:
- Biophysical Chemistry
- Structural Biology
- Analytical Chemistry
Background:
- Understanding biomolecule behavior with temperature is vital for assessing biological activity.
- Optical spectroscopy methods for monitoring heat-induced changes have limitations in interpreting conformational transitions.
- Bovine serum albumin (BSA) is a model protein for studying thermal denaturation.
Purpose of the Study:
- To investigate the structural transitions of bovine serum albumin (BSA) as a function of solution temperature.
- To utilize a temperature-controlled nanoelectrospray ionization (nESI) coupled to trapped ion mobility spectrometry-mass spectrometry (TIMS-MS) for detailed analysis.
- To characterize the unfolding transition and identify distinct structural changes in BSA.
Main Methods:
- Temperature-controlled nanoelectrospray ionization (nESI) coupled to trapped ion mobility spectrometry-mass spectrometry (TIMS-MS).
- Analysis of bovine serum albumin (BSA) in aqueous ammonium acetate solution across a temperature range of approximately 23-70 °C.
- Monitoring changes in charge state distribution and collision cross section (CCS) as indicators of structural transitions.
Main Results:
- BSA exhibited a shift from a narrow, native-like charge state and mobility distribution at lower temperatures to a broad, unfolded-like distribution at higher temperatures.
- While average charge state and CCS suggested a two-state unfolding transition (Tm ≈ 56 °C), detailed CCS profiles revealed at least six distinct structural transitions.
- Nonspecific BSA dimers and trimers were detected at higher concentrations, dissociating early (Td ≈ 34 °C) and potentially influencing monomer melting curves.
Conclusions:
- TIMS-MS coupled with temperature control provides a high-resolution view of protein structural landscapes, revealing complex transitions beyond simple two-state models.
- The method allows for the detection and characterization of early-dissociating oligomers that can affect denaturation studies.
- This technology offers a comprehensive approach to studying solution-phase protein structural dynamics as a function of temperature.
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