The effects of pressure on the energy landscape of proteins

Abstract

Protein dynamics is characterized by fluctuations among different conformational substates, i.e. the different minima of their energy landscape. At temperatures above ~200 K, these fluctuations lead to a steep increase in the thermal dependence of all dynamical properties, phenomenon known as Protein Dynamical Transition. In spite of the intense studies, little is known about the effects of pressure on these processes, investigated mostly near room temperature. We studied by neutron scattering the dynamics of myoglobin in a wide temperature and pressure range. Our results show that high pressure reduces protein motions, but does not affect the onset temperature for the Protein Dynamical Transition, indicating that the energy differences and barriers among conformational substates do not change with pressure. Instead, high pressure values strongly reduce the average structural differences between the accessible conformational substates, thus increasing the roughness of the free energy landscape of the system.

Dynamical properties of myoglobin in an ultraviscous water-glycerol solvent investigated with elastic neutron scattering and FTIR spectroscopy

“…Myoglobin was first dissolved in D 2 O to allow the deuterium exchange of labile hydrogens, and then lyophilized after two days equilibration at room temperature. Then, following a previously reported procedure [26,43], the sample was prepared by successive slow additions of deuterated protein (up to 400 mg) in 400 mg of solvent (66% D 8 -Glycerol -33% D 2 O). Further addition of small amounts D 2 O was necessary to facilitate protein dissolution and excess D 2 O was removed by nitrogen flux upon the sample, until the desired weight proportions were reached: Myoglobin 50%, D 8 -Glycerol 33.5%, D 2 0 16.5%.…”

“…[26]. The total MSD here reported were obtained in the framework of the Gaussian approximation [29], by considering, for each temperature value, a suitable q-range, obtained by fixing q max at the maximum value which consistently satisfies the condition (6〈Δx -46].…”

“…The total MSD here reported were obtained in the framework of the Gaussian approximation [29], by considering, for each temperature value, a suitable q-range, obtained by fixing q max at the maximum value which consistently satisfies the condition (6〈Δx -46]. This allows to maximize the q-range used in each linear fit and therefore to minimize the error in the obtained parameters, by means of an unbiased automatized procedure [26].…”

“…the crossover observed at~180 K in the thermal behavior of protein dynamics, is usually referred to as Protein Dynamical Transition (PDT) [4]. It is still extensively studied because many aspects are not yet clearly understood, as for instance its dependence on the properties of hydration water [17][18][19][20][21], on the characteristics of the external medium in general [22][23][24][25], and on other thermodynamic variables such as pressure [26][27][28].…”

“…In view of the above considerations, we have studied the thermal dependence of the Amide I′ and Amide II′ bands of myoglobin in the temperature range 20-300 K and compared it with the thermal dependence of hydrogen atoms MSD measured in an identical sample with ENS [26]. Aim of the study is to compare the dynamic properties (in particular concerning the PDT) of the protein backbone with those of the protein amino acid side chains.…”