| 摘要: |
Scorpion toxins affecting K+ channels (KTxs) represent important pharmacological tools and potential drug candidates. Here, we report molecular characterization of seven new KTxs in the scorpion Mesobuthus eupeus by cDNA cloning combined with biochemical approaches. Comparative modeling supports that all these KTxs share a conserved cysteine-stabilized alpha-helix/beta-sheet (CSalpha-beta) structural motif despite the differences in protein sequence and size. We investigated functional diversification of two orthologous alpha-KTxs (MeuTXKalpha1 from M. eupeus and BmP01 from M. martensii) by comparing their K+ channel-blocking activities. Pharmacologically, MeuTXKalpha1 selectively blocked Kv1.3 channel with nanomolar affinity (IC50, 2.36 +/- 0.9 nM) whereas only 35% Kv1.1 currents were inhibited at 3 micormolar concentration, showing more than 1271-fold selectivity for Kv1.3 over Kv1.1. This peptide displayed weak effect on Drosophila Shaker channel and no activity on Kv1.2, Kv1.4, Kv1.5, Kv1.6 and hERG channels. Although BmB01 and MeuTXKalpha1 have similar channel spectrum, their affinity and selectivity for these channels largely varies. In comparison with MeuTXKalpha1, BmP01 only exhibits a sub-micromolar affinity (IC50, 133.72 +/- 10.98 nM) for Kv1.3, showing a 57-fold less activity than MeuTXKalpha1. Moreover, it lacks ability to distinguish between Kv1.1 and Kv1.3. We also found that MeuTXKalpha1 inhibited the proliferation of activated T cells induced by phorbol myristate acetate (PMA) and ionomycin at micromolar concentrations. Our results demonstrate that accelerated evolution drives affinity variations of orthologous alpha-KTxs on Kv channels and indicate that MeuTXKalpha1 is a promising candidate to develop an immune modulation agent for human autoimmune diseases.
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