We showed that the patterns of membrane protein-lipid interactions obtained from both the crystal structures and MD trajectories were highly correlated with each other (Figure 1). We also showed that the recognition of lipid tail groups by amino acid residues can be described by the lipophilicity scales (Table 4) and had the same tendency with non-TM proteins (Table 5), while lipid head groups demonstrated considerable variation among individual proteins. We discuss here how our observations can be associated with existing experimental data and previously proposed concepts concerning protein-lipid interactions. We also elaborate on the high propensities of TRP residue for the membrane protein-lipid interface.
Relation of Amino acid propensities to lipid-membrane protein interaction
Since membrane proteins are generally crystallized with detergent molecules used for solubilization and purification, the lipid molecules that remain in the crystal are considered those that are tightly bound to the membrane proteins. On the other hand, the lipid molecules in the first shell, also known as the annular shell around a membrane protein, are in direct contact with the protein and form weak and non-specific interactions according to spin-label EPR and fluorescence quenching experiments [40, 41]. Thus, intuitively, the amino acid propensities from the crystal structures should correspond to propensities for interacting with tightly-bound lipid molecules, while those from the MD trajectories should correspond to propensities for interacting weakly with lipid molecules in the annular shell (although some of these lipid molecules can be tightly bound). It is, therefore, non-trivial that we have observed such a high level of correlation between the propensities derived from these two datasets (Figure 1). Assuming that the tight binding of lipids is achieved by forming a special binding pocket on the surface of a protein, the amino acid composition of such binding pockets appears to be no different from that of other surface positions. This result implies that no special chemical interaction is required for achieving the tight binding of at least the tail portion of lipid molecules, but transmembrane helix packing may create a specific binding pocket for specific lipid types for the protein's function.
Experimental studies of the potassium channel KcsA [4, 42] suggest that the tightly-bound lipids can be essential for its stability and function. The amino acid residues that interact with these tightly-bound lipids must have been selected during the course of evolution. However, our results suggest that these amino acids have been selected not necessarily based on their ability to form special chemical interactions with lipid tails but rather, they are general lipid-binding surface amino acids and happened to have been utilized for offering a physical basis of strong interaction.
For the head group contacts, although the TM and non-TM datasets produced a similar trend (Table 5), a weaker correlation was observed between the propensities derived from the crystal structure and MD datasets than that for the tail group contacts (Figure 1). The difference between the head and tail contacts may be attributable to the larger standard error for the propensities for the head contacts (Table 3). The propensity values were computed for each protein and then averaged and thus, the larger standard error indicates a larger variance among the propensity values derived from different proteins. Indeed, a variety of modes of interaction have been observed between the protein and lipid head groups in our dataset. Head groups of lipids often show disorder in high-resolution X-ray structures even when their tail groups are observed [40, 43]. In our dataset, the head groups of tightly-bound lipids were completely or mostly disordered in rhodopsin (1gzm_A), sensory rhodopsin (1xio_A), succinate:ubiquinone oxidoreductase SQR (2h89_C) and halorhodopsin (3a7k_A); and fully or partially observed but not forming any hydrogen bond in bacteriorhodopsin (1x0i_1), SQR (1zoy_D), V-Type Na+-ATPase (2bl2_I) and ligand-gated ion channel GLIC (3eam_C). In other cases, the head groups appeared and formed hydrogen bonds, while the tail groups were disordered in Ca2+-ATPase (2eau_A), rhomboid protease GlpG (2irv_B), potassium channel Kir (2wll_D) and nitrate reductase A NarGHI (3egw_C).
Experimental studies have shown that differences in the chemical composition of the lipid head group affect the stability and function of membrane proteins, including KcsA, MscL, Ca2+-ATPase and others. Considering all these observations, the role of lipid head-protein interactions is likely to vary among different types of membrane proteins and this notion is consistent with the head contact propensities obtained in this paper, which were diverse and more complex than the tail contact propensities.
Concentration of TRP at a lipid-water interface for anchoring the protein to the membrane
In both the crystal structure and MD datasets, we observed a conspicuously high propensity of TRP residues for contacting lipid molecules (Figure 1), indicating that TRP favors positions in a membrane protein that allow interaction with lipids.
Although TRP is generally not an abundant residue, either in membrane or soluble proteins , TRP has been reported to occur frequently near the membrane boundaries [44–46], as confirmed by our recent statistical analysis . Systematic experimental studies using model peptides and proteins have also produced a similar picture [47–50]. (See Killian and von Hejine  for a review and examples of high-resolution structures are found in Lee .)
The amphiphilic nature of TRP (and also TYR) residues explains why TRP favors to locate at a water-lipid interface; these amphiphilic residues are thought to be locking the membrane protein into the correct location and orientation like anchors or floats at the membrane-water interface. Sansom and colleagues have observed the interfacial anchoring behavior of the amphiphilic residues in their MD simulations of both the outer membrane protein OmpA and the potassium channel KcsA .
All indications are that the significantly high propensities in Figure 1 were obtained as a consequence of the combined effect of the general low abundance and the amphiphilic nature of TRP.