Engineering a Stable and Selective Peptide Blocker of the Kv1.3 Channel in T Lymphocytes Academic Article uri icon

abstract

  • Kv1.3 potassium channels maintain the membrane potential of effector memory (T(EM)) T cells that are important mediators of multiple sclerosis, type 1 diabetes mellitus, and rheumatoid arthritis. The polypeptide ShK-170 (ShK-L5), containing an N-terminal phosphotyrosine extension of the Stichodactyla helianthus ShK toxin, is a potent and selective blocker of these channels. However, a stability study of ShK-170 showed minor pH-related hydrolysis and oxidation byproducts that were exacerbated by increasing temperatures. We therefore engineered a series of analogs to minimize the formation of these byproducts. The analog with the greatest stability, ShK-192, contains a nonhydrolyzable phosphotyrosine surrogate, a methionine isostere, and a C-terminal amide. ShK-192 shows the same overall fold as ShK, and there is no evidence of any interaction between the N-terminal adduct and the rest of the peptide. The docking configuration of ShK-192 in Kv1.3 shows the N-terminal para-phosphonophenylalanine group lying at the junction of two channel monomers to form a salt bridge with Lys(411) of the channel. ShK-192 blocks Kv1.3 with an IC(50) of 140 pM and exhibits greater than 100-fold selectivity over closely related channels. After a single subcutaneous injection of 100 microg/kg, approximately 100 to 200 pM concentrations of active peptide is detectable in the blood of Lewis rats 24, 48, and 72 h after the injection. ShK-192 effectively inhibits the proliferation of T(EM) cells and suppresses delayed type hypersensitivity when administered at 10 or 100 microg/kg by subcutaneous injection once daily. ShK-192 has potential as a therapeutic for autoimmune diseases mediated by T(EM) cells.

authors

  • Pennington, MW
  • Beeton, C
  • Galea, CA
  • Smith, BJ
  • Chi, V
  • Monaghan, KP
  • Garcia, A
  • Rangaraju, S
  • Giuffrida, A
  • Plank, D
  • Crossley, G
  • Nugent, D
  • Khaytin, I
  • LeFievre, Y
  • Peshenko, I
  • Dixon, C
  • Chauhan, S
  • Orzel, A
  • Inoue, T
  • Hu, X
  • Moore, RV
  • Norton, RS
  • Chandy, KG

publication date

  • April 2009