Abstract
Current models of voltage-activated K+ channels predict that the channels are formed by the coassembly of four polypeptide monomers, each of which consists of six transmembrane segments (S1–S6) and long terminal domains. The aqueous pores are thought to be composed of the conserved H-5 regions contributed by four monomers. In this study, two putative membrane-embedded segments of the Shaker K+ channel were synthesized. One segment corresponds to the putative, transmembrane helix S-2 (amino acids 275–300), and the other corresponds to the highly conserved 12 amino acid residues within the H-5 region [amino acids 432–443, designated (12)H-5)]. Structural and functional characterization at elevated lipid/peptide molar ratios (>3000:1) was performed on the two segments, as well as on a previously synthesized 21 amino acid long peptide with a sequence resemblimg the entire H-5 region (designated (21)H-5) (Peled & Shai, 1993). Circular dichroism spectroscopy revealed that S-2 adopts predominantly α-helical structure in both trifluoroethanol and 35 mM SDS (78% or 99%, respectively), while (12)H-5 and (21)H-5 adopt low α-helical structure only in the presence of 35 mM SDS. Functional characterization demonstrated that S-2 and (12)H-5 segments bind to zwitterionic phospholipids, with partition coefficients on the order of 104 M−1. Resonance energy transfer measurements, between donor/acceptor-labeled pairs of peptides, revealed that the peptides self-associate in their membrane-bound state, which may correlate with the existence of functional interactions between the conserved (12)H-5 regions of different subunits of K+ channels (Kirsch et al., 1993). Furthermore, membrane-bound (12)H-5 or (21)H-5 associates with membrane-bound S-2, but does not associate with unrelated, membrane-bound α-helical peptides. These results demonstrate molecular recognition between transmembrane segments of the Shaker K+ channel that might contribute to the oligomerization and correct assembly of the monomers, which result in the formation of a functional channel.
Original language | English |
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Pages (from-to) | 7211-7219 |
Number of pages | 9 |
Journal | Biochemistry |
Volume | 33 |
Issue number | 23 |
DOIs | |
Publication status | Published - 1 Jun 1994 |
All Science Journal Classification (ASJC) codes
- Biochemistry