title: ShK-Dap22, a potent Kv1.3-specific immunosuppressive polypeptide.
abstract: The voltage-gated potassium channel in T lymphocytes, Kv1.3, is an important molecular target for immunosuppressive agents. A structurally defined polypeptide, ShK, from the sea anemone Stichodactyla helianthus inhibited Kv1.3 potently and also blocked Kv1.1, Kv1.4, and Kv1.6 at subnanomolar concentrations. Using mutant cycle analysis in conjunction with complementary mutagenesis of ShK and Kv1.3, and utilizing the structure of ShK, we determined a likely docking configuration for this peptide in the channel. Based upon this topological information, we replaced the critical Lys22 in ShK with the positively charged, non-natural amino acid diaminopropionic acid (ShK-Dap22) and generated a highly selective and potent blocker of the T-lymphocyte channel. ShK-Dap22, at subnanomolar concentrations, suppressed anti-CD3 induced human T-lymphocyte [3H]thymidine incorporation in vitro. Toxicity with this mutant peptide was low in a rodent model, with a median paralytic dose of approximately 200 mg/kg body weight following intravenous administration. The overall structure of ShK-Dap22 in solution, as determined from NMR data, is similar to that of native ShK toxin, but there are some differences in the residues involved in potassium channel binding. Based on these results, we propose that ShK-Dap22 or a structural analogue may have use as an immunosuppressant for the prevention of graft rejection and for the treatment of autoimmune diseases.
authors.affiliation: Departments of Physiology & Biophysics, and Microbiology and Molecular Genetics, University of California, Irvine, California 92697-4560, USA.
authors.forename: K
authors.initials: K
authors.lastname: Kalman
authors.forename: M W
authors.initials: MW
authors.lastname: Pennington
authors.forename: M D
authors.initials: MD
authors.lastname: Lanigan
authors.forename: A
authors.initials: A
authors.lastname: Nguyen
authors.forename: H
authors.initials: H
authors.lastname: Rauer
authors.forename: V
authors.initials: V
authors.lastname: Mahnir
authors.forename: K
authors.initials: K
authors.lastname: Paschetto
authors.forename: W R
authors.initials: WR
authors.lastname: Kem
authors.forename: S
authors.initials: S
authors.lastname: Grissmer
authors.forename: G A
authors.initials: GA
authors.lastname: Gutman
authors.forename: E P
authors.initials: EP
authors.lastname: Christian
authors.forename: M D
authors.initials: MD
authors.lastname: Cahalan
authors.forename: R S
authors.initials: RS
authors.lastname: Norton
authors.forename: K G
authors.initials: KG
authors.lastname: Chandy
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006801:Humans; D007166:Immunosuppressive Agents / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D051664:Kv1.3 Potassium Channel; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D015221:Potassium Channels / Q000378:metabolism*; D024642:Potassium Channels, Voltage-Gated; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism
chemical_list: D007166:Immunosuppressive Agents; C496973:KCNA3 protein, human; C496974:Kcna3 protein, mouse; D051664:Kv1.3 Potassium Channel; D010455:Peptides; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011994:Recombinant Proteins