1C2U

SOLUTION STRUCTURE OF [ABU3,35]SHK12-28,17-32

1C2U の概要

• エントリーDOI: 10.2210/pdb1c2u/pdb
• 分子名称: SYNTHETIC PEPTIDE ANALOGUE OF SHK TOXIN (1 entity in total)
• 機能のキーワード: shk toxin, potassium channel, disulphide bonds, analogues, structure-function, solution structure, toxin
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 3973.70

構造登録者

Pennington, M.W.,Lanigan, M.D.,Kalman, K.,Manhir, V.M.,Rauer, H.,McVaugh, C.T.,Behm, D.,Donaldson, D.,Chandy, K.G.,Kem, W.R.,Norton, R.S. (登録日: 1999-07-27, 公開日: 1999-11-10, 最終更新日: 2021-11-03)

主引用文献

Pennington, M.W.,Lanigan, M.D.,Kalman, K.,Mahnir, V.M.,Rauer, H.,McVaugh, C.T.,Behm, D.,Donaldson, D.,Chandy, K.G.,Kem, W.R.,Norton, R.S.
Role of disulfide bonds in the structure and potassium channel blocking activity of ShK toxin.
Biochemistry, 38:14549-14558, 1999

PubMed Abstract: ShK toxin, a potassium channel blocker from the sea anemone Stichodactyla helianthus, is a 35 residue polypeptide cross-linked by three disulfide bridges: Cys3-Cys35, Cys12-Cys28, and Cys17-Cys32. To investigate the role of these disulfides in the structure and channel-blocking activity of ShK toxin, a series of analogues was synthesized by selective replacement of each pair of half-cystines with two alpha-amino-butyrate (Abu) residues. The remaining two disulfide pairs were formed unambiguously using an orthogonal protecting group strategy of Cys(Trt) or Cys(Acm) at the appropriate position. The peptides were tested in vitro for their ability to block Kv1.1 and Kv1.3 potassium channels and their ability to displace [(125)I]dendrotoxin binding to rat brain synaptosomal membranes. The monocyclic peptides showed no activity in these assays. Of the dicyclic peptides, [Abu12,28]ShK(3-35,17)(-)(32) (where the subscript indicates disulfide connectivities) had weak activity on Kv1.3 and Kv1.1. [Abu17,32]ShK(3-35,12)(-)(28) blocked Kv1.3 with low nanomolar potency, but was less effective (being comparable to [Abu12,28]ShK(3-35,17)(-)(32)) against Kv1.1. [Abu3, 35]ShK(12-28,17)(-)(32), retained high picomolar affinity against both channels. Corroborating these results, [Abu3,35]ShK(12-28, 17)(-)(32) had an IC(50) ratio relative to native toxin of 18 in the displacement assay, whereas [Abu17,32]ShK(3-35,12)(-)(28) and [Abu12, 28]ShK(3-35,17)(-)(32) had ratios of 69 and 390, respectively. Thus, the disulfide bond linking the N- and C-terminal regions is less important for activity than the internal disulfides. NMR analysis of the [Abu12,28] and [Abu17,32] analogues indicated that they had little residual structure, consistent with their significantly reduced activities. By contrast, [Abu3,35]ShK(12-28,17)(-)(32) had a moderately well-defined solution structure, with a mean pairwise root-mean-square deviation of 1.33 A over the backbone heavy atoms. This structure nevertheless showed significant differences from that of native ShK toxin. The possible interactions of this analogue with the channel and the distinction between native secondary and tertiary structure on one hand and global topology imposed by the disulfide bridges on the other are discussed.

PubMed: 10545177
DOI: 10.1021/bi991282m

実験手法

SOLUTION NMR