Properties of the protein matrix revealed by the free energy of cavity formation.
BACKGROUND
The classical picture of the hydrophobic stabilization of proteins invokes a resemblance between the protein interior and nonpolar solvents, but the extent to which this is the case has often been questioned. The protein interior is believed to be at least as tightly packed as organic crystals, and was shown to have very low compressibility. There is also evidence that these properties are not uniform throughout the protein, and conflicting views exist on the nature of sidechain packing and on its influence on the properties of the protein.


RESULTS
In order to probe the physical properties of the protein, the free energy associated with the formation of empty cavities has been evaluated for two proteins: barnase and T4 lysozyme. To this end, the likelihood of encountering such cavities was computed from room temperature molecular dynamics trajectories of these proteins in water. The free energy was evaluated in each protein taken as a whole and in submolecular regions. The computed free energies yielded information on the manner in which empty space is distributed in the system, while the latter undergoes thermal motion, a property hitherto not analyzed in heterogeneous media such as proteins. Our results showed that the free energy of cavity formation is higher in proteins than in both water and hexane, providing direct evidence that the native protein medium differs in fundamental ways from the two liquids. Furthermore, although the packing density was found to be higher in nonpolar regions of the protein than in polar ones, the free energy cost of forming atomic size cavities is significantly lower in nonpolar regions, implying that these regions contain larger chunks of empty space, thereby increasing the likelihood of containing atomic size packing defects. These larger empty spaces occur preferentially where buried hydrophobic sidechains belonging to secondary structures meet one another. These particular locations also appear to be more compressible than other parts of the core or surface of the protein.


CONCLUSIONS
The cavity free energy calculations described here provide a much more detailed physical picture of the protein matrix than volume and packing calculations. According to this picture, the packing of hydrophobic sidechains is tight in the interior of the protein, but far from uniform. In particular, the packing is tighter in regions where the backbone forms less regular hydrogen-bonding interactions than at interfaces between secondary structure elements, where such interactions are fully developed. This may have important implications on the role of sidechain packing in protein folding and stability.
Kocher
J P
JP
Unité de Conformation de Macromolécules Biologiques, Université Libre de Bruxelles, Belgium.
Prévost
M
M
Wodak
S J
SJ
Lee
B
B
D001426:Bacterial Proteins; D017122:Bacteriophage T4 / Q000201:enzymology; D004795:Enzyme Stability; D008958:Models, Molecular; D009113:Muramidase / Q000737:chemistry*; D009154:Mutation; D011487:Protein Conformation; D017510:Protein Folding; D012260:Ribonucleases / Q000737:chemistry*; D012997:Solvents / Q000494:pharmacology; D013816:Thermodynamics
D001426:Bacterial Proteins; D012997:Solvents; D012260:Ribonucleases; C019760:Bacillus amyloliquefaciens ribonuclease; D009113:Muramidase