GENETICS OF TRANSMEMBRANE SEGMENT INTERACTION

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Hydrophobic transmembrane (TM's) of membrane proteins serve various roles: 1) assembly of individual membrane proteins into their tertiary structures, 2) homodimerization or homo-oligomerization of individual membrane proteins or 3) assembly of hetero-oligomeric membrane protein complexes. The best characterized example of TM interactions is the single homodimerizing TM of human erythrocyte glycophorin A. This proposal is designed to detect and genetically characterize pairs of interacting TM segments from proteins of the bacterium E. coli. Any newly identified dimerizing TM's will be analyzed in collaboration with co-investigators by theoretical and biophysical approaches to determine and understand the structure and nature of the TM interactions. Mutational studies will be carried out to define amino acids critical to TM dimerization. Ultimately, the results could allow predications of aspects of the tertiary structure of membrane proteins and membrane protein complexes. A genetic system was developed that can be used to detect those TM segments that homo-dimerize (and probably oligomerize). Such TM's are detected by their ability to dimerize the "head-piece" of bacteriophage lambda repressor, thus allowing repression of a lambda cI-phage entering the bacterial cell. A screen of the E. coli genome will be done for portions of genes coding entering the bacterial cell. A screen of the E. coli genome will be done for portions of genes coding for TM's that homodimerize. A system will be developed for the detection of dimerization of heterologous pairs of TM's. The possibility that homo- or hetero-oligomerization of entire membrane proteins may be assayed by this approach will also be tested. The possibility can be tested with already known pairs of interacting membrane proteins. Such studies could allow us to further define, within such pairs of proteins, those TM's responsible for the assembly of the complexes. This latter approach could provide a general assay for defining membrane protein complexes. The overall project would add to our catalogue of structures required for interactions between TM's.