Lipid-helix interactions in membrane receptors.

The integration of the transmembrane (TM) proteins in the lipid bilayer depend on lipid-protein interactions. The few crystal structures known for multiple membrane spanning proteins suggest a basic form composed of bundles of a-helices that pack together. In the membrane helix-helix and helix-lipid interaction define the helix arrangement in the folded receptor. Thus, the lipid-protein interaction may be used to gain information on the structure of membrane proteins integrated in the lipid bilayer. Also, i t has been shown that only correctly determined lipid-exposed faces of TM helices of Bacteriorhodopsin led to the successful prediction of helix packing in Bacteriorhodopsin [ I ] . The conformations of the domains and subunits of high affinity IgE receptor were studied by CD, NMR and molecular modelling and the structure of the subunit was proposed [2]. Here we report molecular mechanics and dynamics studies lipid-TM helix hydrophobic interaction sites of two TM proteins: the high affinity IgE receptor and Bacteriorhodopsin, as a test case. The interaction energies between each TM helix and the dodecane molecule were calculated by X-PLOR using the CHARMM 22x parameter set, and dielectric constant (E) of 1.0 (dodecane was used instead of lipid to avoid electrostatic and polar interactions). Helical backbone geometry was explicitly maintained during simulations through imposition of a one-sided distance restraint with an upper boundary of 3.2A between I oxygens and 1+4 nitrogens along the length of the helix. The TM helix and dodecane were always oriented with a long axis parallel to each other. The eight positions were considered, with separation of a 5A between the helix and dodecane (Figure 1 .). The modified simulated annealing protocol was used [3], namely, 1) a 2ps molecular dynamics heating stage at 600K with a timestep of Ifs, 2) a 2ps constant temperature molecular dynamics simulation at 300k with timestep of 2fs. and 3) 1000 steps of conjugate gradient minimisation. The a-helical hydrogen bond restraints were active during simulation stages, but no other restraints were applied during constant temperature simulation and energy minimisation stages. The interaction energies between dodecane and TM helices were calculated relative to those when the two molecules were far apart: GE=E,,,,,,,-E,,,. Thus, negative values of interaction energies corresponded to stabilization. All energies in Table 1. (for Bacteriorhodopsin) and 2. (for the high affinity IgE receptor) were negative. The lowest energy positions indicated favourable lipid-hydrocarbon binding sites. The energy differences between lowest and highest energy positions were in the range from 0 to 18 kcal/mol. The essential focus of the interaction between the dodecane and helix included packing interactions involving the hydrophobic side-chains in the protein. Usually, two or three hydrocarbon interaction sites with lower energies were detected on the transmembrane helices as a function of rotation through 360" of dodecane around the helical axis. In most of cases, dodecane was moving towards the binding sites 3f the TM helices