Membrane interaction and self-assembly within phospholipid membranes of synthetic segments corresponding to the H-5 region of the shaker K+ channel.

The voltage-activated K+ channels are assumed to be formed by the coassembly of four polypeptide monomers. Each of these monomers is postulated to consist of six transmembrane segments (S1 to S6), and long N- and C-terminal domains. The highly conserved linker, H-5, between the fifth and the sixth transmembrane segments, is hypothesized to line the lumen of the K+ channel formed by the bundle of the transmembrane segments of the monomers. Herein we utilize the spectrofluorometric approach and investigate the interaction with phospholipid membranes of fluorescently-labeled synthetic peptides, whose sequences are derived from the H-5 region. Binding experiments reveal that the peptides can strongly bind to phospholipid membranes with partition coefficients on the order of 10(4) M-1. However, a truncated peptide without four amino acids within the most conserved region (amino acids 432-435) did not bind to the membranes at all. Moreover, the single substitution of a conserved tryptophan at position 435 to serine reduced the partition coefficient of the peptide approximately 5-fold, which may account for a mutated K+ channel with this substitution not producing functional channels (Yool & Schwarz, 1991). Structural characterization using circular dichroism spectroscopy (CD) reveals that H-5 can partially adopt an alpha-helix structure in hydrophobic environments. Resonance energy transfer (RET) experiments reveal that the H-5-derived segments can self-assemble within the membrane but cannot coassemble with other unrelated membrane-bound peptides. The results herein support the hypothesis that H-5 segments are packed in close proximity and might participate in mediating the appropriate assembly of the core region of K+ channel monomers.