While several reports have suggested a role for helix-helix interactions in membrane protein oligomerization, there are few direct biochemical data bearing on this subject. Here, using mutational analysis, the dimerization of the transmembrane alpha-helix of giycophorin A in a detergent environment is shown to be both spontaneous and highly specific. Very subtle changes in the side-chain structure at certain sensitive positions disrupt the helix-helix association. These sensitive positions occur at approximately every 3.9 residues along the helix, consistent with their comprising the interface of a closely fit transmembraneous supercoil of alpha-helices. Simultaneous modeling studies of the helix dimer using simulated annealing techniques are in excellent agreement with the experimental data, and predict a pair of alpha-helices in a right-handed supercoil. By contrast with other examples of transmembrane alpha-helix association, the set of interfacial residues in this case contains no highly polar groups. Amino acids with aliphatic side-chains define much of the interface, indicating that precise packing interactions between the helices may provide much of the energy for association. Possible general implications of this result for the oligomerization of other transmembrane alpha-helices are discussed.

The interfacial residues predicted by the above studies comprise the sequence LIXXGVXXGVXXT. Placement of this pattern in the context of poly-L-leucine results in a transmembrane alpha-helix which forms dimers in SDS, and interacts specifically with the transmembrane domain of glycophorin. Mutational analysis and modeling studies indicate that the interface of this dimer is identical to that seen for glycophorin. This sequence pattern has also been introduced into the single transmembrane domain of both the neu oncogene product and the epidermal growth factor receptor, using the smallest possible number of mutations. In both cases, the resultant transmembrane domains form dimers and will interact with the transmembrane domain of glycophorin. Studies are described of the effects of these mutations upon the activity of the receptor molecules in vivo.