The behavior of the hydrophobic effect under pressure and protein denaturation

Abstract

It is well known that proteins denature under high pressure. The mechanism that underlies such a process is still not clearly understood, however, giving way to controversial interpretations. Using molecular dynamics simulation on systems that may be regarded experimentally as limiting examples of the effect of high pressure on globular proteins, such as lysozyme and apomyoglobin, we have effectively reproduced such similarities and differences in behavior as are interpreted from experiment. From the analysis of such data, we explain the experimental evidence at hand through the effect of pressure on the change of water structure, and hence the weakening of the hydrophobic effect that is known to be the main driving force in protein folding.

Figure 1

Hydrogen bond distribution of SPC/E water at 300 K at: 1 bar (black), 3 kbar (dark gray), and 10 Kbar (light gray), as obtained by MD simulation. We can observe an increase in three and five HB/mol coordination states, at the expense of four HB/mol, with the rise in pressure. The criteria used to consider the formation of a hydrogen bond was an OHO angle along the bond not <145°, and an O–H distance not >0.24 nm. The error bars have not been incorporated because they are undetectable visually.

Figure 2

Normalized RMSD (average over all α-carbon atoms of the protein during the whole simulation). The RMSD of the position of α-carbon atoms of the protein along the simulation time shows that at 1 bar the protein remains fluctuating around a homogeneous structure, whereas at 3 kbar a monotonic deviation from the initial structure may be observed. For a better comparison of the high and low pressure RMSD, data has been normalized with the standard deviation values on the ns scale for each system respectively.

Figure 3

(a) Hydrophobic and (b) hydrophilic SAS: 1 bar (solid line); 3 kbar (dotted line). The horizontal lines (– · – ·) correspond to 1 bar average. We can see that both surfaces at 1 bar fluctuate around a relatively stable value, but the 3 kbar surfaces do not. Although the 3 kbar hydrophilic surface behavior does not allow for any definite conclusion, the constant increase rate in the hydrophobic SAS along the total run shows the wakening of the hydrophobic interaction, which in turn leads to protein unfolding.

Figure 4

Connelly surface of apomyoglobin. (a) 1 bar. (b) 3 kbar. The color surface corresponds to the electrostatic potential: gray charged, white neutral. This surface is computed rolling a sphere of radius equal to that of a water molecule along the protein, thus evaluating the molecular area accessible to the solvent.