Subunit Composition of Brain Voltage-gated Potassium Channels Determined by Hongotoxin-1, a Novel Peptide Derived fromCentruroides limbatus Venom
| Autor: | Gregory J. Kaczorowski, Jörg Mitterdorfer, Maria L. Garcia, Hans-Günther Knaus, Randal M. Bugianesi, Alexandra Koschak |
|---|---|
| Rok vydání: | 1998 |
| Předmět: |
Potassium Channels
Protein Conformation Immunoprecipitation Protein subunit Molecular Sequence Data Neurotoxins Scorpion Venoms Peptide Biology Ligands complex mixtures Biochemistry Animals natural sciences Amino Acid Sequence Receptor Molecular Biology Peptide sequence Brain Chemistry chemistry.chemical_classification Sequence Homology Amino Acid urogenital system Cell Biology Transfection Voltage-gated potassium channel Recombinant Proteins Rats Membrane nervous system chemistry Shaker Superfamily of Potassium Channels Synaptic Vesicles biological phenomena cell phenomena and immunity Ion Channel Gating Sequence Analysis Protein Binding |
| Zdroj: | Journal of Biological Chemistry. 273:2639-2644 |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.273.5.2639 |
| Popis: | Five novel peptidyl inhibitors of Shaker-type (Kv1) K+ channels have been purified to homogeneity from venom of the scorpion Centruroides limbatus. The complete primary amino acid sequence of the major component, hongotoxin-1 (HgTX1), has been determined and confirmed after expression of the peptide in Escherichia coli. HgTX1 inhibits 125I-margatoxin binding to rat brain membranes as well as depolarization-induced 86Rb+ flux through homotetrameric Kv1.1, Kv1. 2, and Kv1.3 channels stably transfected in HEK-293 cells, but it displays much lower affinity for Kv1.6 channels. A HgTX1 double mutant (HgTX1-A19Y/Y37F) was constructed to allow high specific activity iodination of the peptide. HgTX1-A19Y/Y37F and monoiodinated HgTX1-A19Y/Y37F are equally potent in inhibiting 125I-margatoxin binding to rat brain membranes as HgTX1 (IC50 values approximately 0.3 pM). 125I-HgTX1-A19Y/Y37F binds with subpicomolar affinities to membranes derived from HEK-293 cells expressing homotetrameric Kv1.1, Kv1.2, and Kv1.3 channels and to rat brain membranes (Kd values 0.1-0.25 pM, respectively) but with lower affinity to Kv1.6 channels (Kd 9.6 pM), and it does not interact with either Kv1.4 or Kv1.5 channels. Several subpopulations of native Kv1 subunit oligomers that contribute to the rat brain HgTX1 receptor have been deduced by immunoprecipitation experiments using antibodies specific for Kv1 subunits. HgTX1 represents a novel and useful tool with which to investigate subclasses of voltage-gated K+ channels and Kv1 subunit assembly in different tissues. |
| Databáze: | OpenAIRE |
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