Hydration-State Change of Horse Heart Cytochrome c Corresponding to Trifluoroacetic-Acid-Induced Unfolding

Abstract

We investigate the hydration state of horse-heart cytochrome c (hh cyt c) in the unfolding process induced by trifluoroacetic acid (TFA). The conformation of hh cyt c changes from the native (N) state (2.9 < pH < 6.0) to the acid-unfolded (U(A)) state (1.7 < pH < 2.0) to the acid-induced molten globule (A) state (pH ∼1.2). Hydration properties of hh cyt c during this process are measured at 20°C by high-resolution dielectric relaxation (DR) spectroscopy, UV-vis absorbance, and circular dichroism spectroscopy. Constrained water of hh cyt c is observed at every pH as an ∼5-GHz Debye component (DC) (DR time, τ(D) ∼30 ps) and its DR amplitude (DRA) is increased by 77% upon N-to-U(A) transition, when pH changes from 6.0 to 2.0. Even in the N state, the DRA of the constrained-water component is found to be increased by 22% with decreasing pH from 6.0 to 2.9, suggesting an increase in the accessible surface area of native hh cyt c. Moreover, hypermobile water around native hh cyt c is detected at pH 6.0 as a 19-GHz DC (τ(D) ∼ 8.4 ps <τ(DW) = 9.4 ps), but is not found at other pH values. The DRA signal of constrained water is found to return to the pH 2.9 (N-state) level upon U(A)-to-A transition. Fast-response water (slightly slower than bulk) around A-state hh cyt c is detected at pH 1.2, and this suggests some accumulation of TFA(-) ions around the peptide chain. Thus, this high-resolution DR spectroscopy study reveals that hh cyt c exhibits significant hydration-state change in the TFA-unfolding process.

What is “hypermobile” water?: detected in alkali halide, adenosine phosphate, and F-actin solutions by high-resolution microwave dielectric spectroscopy

“…4 [31]. No hypermobile component was found in the hydration shell of S1 itself as well as many other globular proteins [32,33] except for G-actin [14,31] and horse heart cytochrome c [15]. The results suggest that the solvent space around S1 bound to F-actin is asymmetric in water-mobility.…”

“…Some of those modified γ dispersions of alkali-halide solutions were further analyzed using a series of Debye components in our high-resolution DRS study [9], where volume fraction and DR property of the hydration water in an aqueous solution of salt at low concentration were analyzed by combining mixture theories [10,11] and the Debye component analysis for the dielectric spectra in the frequency range of 0.2−26 GHz. Although dielectric mixture theories [10][11][12] had been constructed for the analyses of colloidal solutions or emulsions, such theories evaluate dielectric property of a spherical/ellipsoidal water region containing a solute, assuming that the continuous bulk-solvent phase exists and suspends solute particles [13][14][15].…”

Physical driving force of actomyosin motility based on the hydration effect

“…1. The protein structure is strongly coupled with the solvation state (Miyashita et al, 2013;Suzuki et al, 1951;Wazawa, Miyazaki, Sambongi, & Suzuki, 2010).…”

“…Here, we summarize the relevant points for this model (see Figure ). The protein structure is strongly coupled with the solvation state (Miyashita et al, ; Suzuki et al, ; Wazawa, Miyazaki, Sambongi, & Suzuki, ). During the fluctuation of the protein structure, the change in the protein solvation free energy (Δμ) is mostly compensated by the change in the protein structure energy (Karino & Matubayasi, ). From points 1 and 2 above, a change in the Δμ of the protein or the protein assembly induces a structural change in the protein or an arrangement change of the protein assembly.From points 1 to 3 above, we propose a driving force hypothesis based on the hydration effects on the protein structures and the following assumptions. When an M.ADP.Pi(c) binds to F‐actin ([a] to [b] in Figure ), the M.ADP.Pi(c) isomerizes to M.ADP.Pi(o) and simultaneously several actin protomers on the back side of M change their structure. This reduces the radial electric field strength and therefore the hyper‐mobile water (HMW; colored in light blue ) in close proximity to F‐actin ([b] to [c]).…”

Quantification of solvent water and hydration dynamic of thermo‐sensitive microgel by dielectric spectroscopy

“…Water is the most important and active molecule in all biomacromolecules, and it is thought to affect the function, structure, stability, and dynamics of other biological macromolecules . Therefore, the quantification of water will undoubtedly promote the understanding of many complex biological mechanisms, such as protein folding .…”