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. Author manuscript; available in PMC: 2023 May 9.
Published in final edited form as: Annu Rev Biophys. 2021 Dec 21;51:63–77. doi: 10.1146/annurev-biophys-102221-101121

Figure 1.

Figure 1

Thermodynamics, electrospray ionization (ESI), and capturing non-native structures. (a) Illustration of the ESI process beginning with formation of nanodroplets. In solution, rapid interconversion of protein structures [native (N), intermediate (I), and denatured (D)] is mediated by solvent and other chemical species, e.g., buffer, pH, temperature, and ligands. During early states of ESI, the droplet contains the solution populations of conformers (bottom). As the droplet dries, the solution equilibrium is quenched by evaporative cooling (middle), and during the late stages of solvent removal, protein structures are kinetically trapped on a new, gas-phase free energy landscape associated with the solvent-free ion (top). T-dependent charge state is shown as a function; water pH is 2.6 and Tm = 48.5 ± 0.3°C, in good agreement with traditional calorimetry studies (34) and mass spectra. (b) Temperature dependence of equilibrium association constants (Ka). Ka can follow linear (negative: green line; positive: purple line) and non-linear (concave: orange line; convex: blue line) trends depending on the sign and magnitude of thermodynamic parameters (ΔH and TΔS). (c) Melting for chymotrypsin inhibitory 2 (CI-2). (d) A plot of relative abundance as a function of solution temperature for each of the six individual charge states of CI-2 (56). For a free-energy landscape depicting ΔG as a function of collision cross section (CCS) (for each charge state) showing 41 unique solution structures, the reader is referred to Reference 56. (Top) T-dependent CCS distributions of wild-type (WT) 6+ through 11+ charge states. Vertical lines represent peak centers for each Gaussian function, and the color represents the corresponding conformational group determined from grouping analysis of the WT and mutant (L32A, A16G, K11A) data. (Bottom) Scaled plots of the relative abundance of representative groups as a function of solution temperature. The arrows indicate the respective Tm or Tf values for each conformer family. Panels a, b, and d adapted with permission from Reference 59, copyright 2021 American Chemical Society. Panel c adapted with permission from Reference 56, copyright 2020 American Chemical Society.