Transmembrane domain mediated self-assembly of major coat protein subunits from Ff bacteriophage.

The 50-residue major coat protein (MCP) of Ff bacteriophage exists as a single-spanning membrane protein in the Escherichia coli host inner membrane prior to assembly into lipid-free virions. Here, the molecular bases for the specificity and stoichiometry that govern the protein-protein interactions of MCP in the host membrane are investigated in detergent micelles. To address these structural issues, as well as to circumvent viability requirements in mutants of the intact protein, peptides corresponding to the effective alpha-helical TM segment of wild-type and mutant bacteriophage MCPs were synthesized. Fluorescence resonance energy transfer (FRET) experiments on the dansyl and dabcyl-labeled MCP TM domain peptides in detergent micelles demonstrated that the peptides specifically associate into non-covalent homodimers, as postulated for the biologically relevant membrane-embedded MCP oligomer. MCP peptides labeled with short-range pyrene fluorophores at the N terminus displayed excimer fluorescence consistent with homodimerization occurring in a parallel fashion. Variant peptides synthesized with single substitutions at helix-interactive positions displayed a wide range of dimer/monomer ratios on SDS-PAGE gels, which are interpreted in terms of steric volume, presence or absence of beta-branching, and the effect of polar substituents. The overall results indicate discrete roles for helix-helix interfacial residues as packing recognition elements in the membrane-inserted state, and suggest a possible correlation between phage viability and efficacy of MCP TM-TM interactions.