Protein dynamics: from rattling in a cage to structural relaxation.

We present an overview of protein dynamics based mostly on results of neutron scattering, dielectric relaxation spectroscopy and molecular dynamics simulations. We identify several major classes of protein motions on the time scale from faster than picoseconds to several microseconds, and discuss the coupling of these processes to solvent dynamics. Our analysis suggests that the microsecond backbone relaxation process might be the main structural relaxation of the protein that defines its glass transition temperature, while faster processes present some localized secondary relaxations. Based on the overview, we formulate a general picture of protein dynamics and discuss the challenges in this field.

Influence of sorbitol on protein crowding in solution and freeze-concentrated phases.

Small-angle neutron scattering was employed to study protein crowding under freezing conditions that mimic those used in pharmaceutical processing. The results demonstrate that, although there is an increase in heterogeneity as the temperature is reduced, sorbitol reduces protein crowding in both solution and freeze-concentrated phases, thus protecting the protein from forming oligomers or irreversible aggregates.

A broad glass transition in hydrated proteins.

We performed Raman and Brillouin scattering measurements to estimate glass transition temperature, T(g), of hydrated protein. The measurements reveal very broad glass transition in hydrated lysozyme with approximate T(g) approximately 180+/-15 K. This result agrees with a broad range of T(g) approximately 160-200 K reported in literature for hydrated globular proteins and stresses the difference between behavior of hydrated biomolecules and simple glass-forming systems. Moreover, the main structural relaxation of the hydrated protein system that freezes at T(g) approximately 180 K remains unknown. We emphasize the difference between the "dynamic transition", known as a sharp rise in mean-squared atomic displacement at temperatures around T(D) approximately 200-230 K, and the glass transition. They have different physical origin and should not be confused.

Dynamics of biological macromolecules: not a simple slaving by hydration water.

We studied the dynamics of hydrated tRNA using neutron and dielectric spectroscopy techniques. A comparison of our results with earlier data reveals that the dynamics of hydrated tRNA is slower and varies more strongly with temperature than the dynamics of hydrated proteins. At the same time, tRNA appears to have faster dynamics than DNA. We demonstrate that a similar difference appears in the dynamics of hydration water for these biomolecules. The results and analysis contradict the traditional view of slaved dynamics, which assumes that the dynamics of biological macromolecules just follows the dynamics of hydration water. Our results demonstrate that the dynamics of biological macromolecules and their hydration water depends strongly on the chemical and three-dimensional structures of the biomolecules. We conclude that the whole concept of slaving dynamics should be reconsidered, and that the mutual influence of biomolecules and their hydration water must be taken into account.

Influence of hydration on protein dynamics: combining dielectric and neutron scattering spectroscopy data.

Combining dielectric spectroscopy and neutron scattering data for hydrated lysozyme powders, we were able to identify several relaxation processes and follow protein dynamics at different hydration levels over a broad frequency and temperature range. We ascribe the main dielectric process to protein's structural relaxation coupled to hydration water and the slowest dielectric process to a larger scale protein's motions. Both relaxations exhibit a smooth, slightly super-Arrhenius temperature dependence between 300 and 180 K. The temperature dependence of the slowest process follows the main dielectric relaxation, emphasizing that the same friction mechanism might control both processes. No signs of a proposed sharp fragile-to-strong crossover at T approximately 220 K are observed in temperature dependences of these processes. Both processes show strong dependence on hydration: the main dielectric process slows down by six orders with a decrease in hydration from h approximately 0.37 (grams of water per grams of protein) to h approximately 0.05. The slowest process shows even stronger dependence on hydration. The third (fastest) dielectric relaxation process has been detected only in samples with high hydration ( h approximately 0.3 and higher). We ascribe it to a secondary relaxation of hydration water. The mechanism of the protein dynamic transition and a general picture of the protein dynamics are discussed.

Nanosecond relaxation dynamics of hydrated proteins: water versus protein contributions.

We have studied picosecond to nanosecond dynamics of hydrated protein powders using dielectric spectroscopy and molecular dynamics (MD) simulations. Our analysis of hydrogen-atom single particle dynamics from MD simulations focused on "main" (τ(main) ≈ tens of picoseconds) and "slow" (τ(slow) ≈ nanosecond) relaxation processes that were observed in dielectric spectra of similar hydrated protein samples. Traditionally, the interpretation of these processes observed in dielectric spectra has been ascribed to the relaxation behavior of hydration water tightly bounded to a protein and not to protein atoms. Detailed analysis of the MD simulations and comparison to dielectric data indicate that the observed relaxation process in the nanosecond time range of hydrated protein spectra is mainly due to protein atoms. The relaxation processes involve the entire structure of protein including atoms in the protein backbone, side chains, and turns. Both surface and buried protein atoms contribute to the slow processes; however, surface atoms demonstrate slightly faster relaxation dynamics. Analysis of the water molecule residence and dipolar relaxation correlation behavior indicates that the hydration water relaxes at much shorter time scales.

Conductivity in hydrated proteins: no signs of the fragile-to-strong crossover.

Dielectric spectroscopy studies of hydrated protein demonstrate smooth temperature variations of conductivity. This observation suggests no cusplike fragile-to-strong crossover (FSC) in the protein's hydration water. The FSC at T approximately 220 K was postulated recently on the basis of neutron scattering studies [Chen, Proc. Natl. Acad. Sci. U.S.A. 103, 9012 (2006)] and was proposed to be the main cause for the dynamic transition in hydrated proteins. Following Swenson et al. , we ascribe the neutron results to a secondary relaxation. We emphasize that no cusplike solvent behavior is required for the protein's dynamic transition.

The origin of the dynamic transition in proteins.

Despite extensive efforts in experimental and computational studies, the microscopic understanding of dynamics of biological macromolecules remains a great challenge. It is known that hydrated proteins, DNA and RNA, exhibit a so-called "dynamic transition." It appears as a sharp rise of their mean-squared atomic displacements r2 at temperatures above 200-230 K. Even after a long history of studies, this sudden activation of biomolecular dynamics remains a puzzle and many contradicting models have been proposed. By combining neutron and dielectric spectroscopy data, we were able to follow protein dynamics over an extremely broad frequency range. Our results show that there is no sudden change in the dynamics of the protein at temperatures around approximately 200-230 K. The protein's relaxation time exhibits a smooth temperature variation over the temperature range of 180-300 K. Thus the experimentally observed sharp rise in r2 is just a result of the protein's structural relaxation reaching the limit of the experimental frequency window. The microscopic mechanism of the protein's structural relaxation remains unclear.