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GaTx2 (gating modifier of anion channels 2) was isolated from the venom of Leiurus quinquestriatus hebraeus. GaTx2 is the most potent peptide inhibitor of ClC-2 (CLCN2) chloride channel ever described. Kd value is close to 20 pM. GaTx2slows ClC-2 activation but without altering channel conductance. The effect is voltage-dependent. This inhibitory effect was highlighted on rabbit ClC-2 channels expressed in oocytes. It has no effect on ClC-0, ClC-1, ClC-3, ClC-4, CFTR, GABAC, Xenopus ClCa, Shaker B or Kv1.2 channels. Structurally, GaTx2 is composed of two β-strands and one α-helix.This peptide is also called Leiuropeptide II. Bears 89, 93 and 96% identity with OdK1, neurotoxin PO1 and leiuropeptide III, respectively.
AA sequence: Val-Ser-Cys3-Glu-Asp-Cys6-Pro-Asp-His-Cys10-Ser-Thr-Gln-Lys-Ala-Arg-Ala-Lys-Cys19-Asp-Asn-Asp-Lys-Cys24-Val-Cys26-Glu-Pro-Ile-OH
Disulfide bonds between: Cys3-Cys19, Cys6-Cys24, and Cys10-Cys26
Length (aa): 29
Formula: C125H199N39O47S6
Molecular Weight: 3191.25 Da
Appearance: White lyophilized solid
Solubility: water and saline buffer
CAS number: not available
Source: Synthetic
Purity rate: > 98 %
Multiple sodium and chloride channels on the apical surface of nasal epithelial cells contribute to periciliary fluid homeostasis, a function that is disrupted in patients with cystic fibrosis (CF). Among these channels is the chloride channel CLCN2, which has been studied as a potential alternative chloride efflux pathway in the absence of CFTR. The object of the present study was to use the nasal potential difference test (NPD) to quantify CLCN2 function in an epithelial-directed TetOn CLCN2 transgenic mouse model (TGN-K18rtTA-hCLCN2) by using the putative CLCN2 pharmacological agonist lubiprostone and peptide inhibitor GaTx2. Lubiprostone significantly increased chloride transport in the CLCN2-overexpressing mice following activation of the transgene by doxycycline. This response to lubiprostone was significantly inhibited by GaTx2 after CLCN2 activation in TGN-CLCN2 mice. Cftr(-/-) and Clc2(-/-) mice showed hyperpolarization indicative of chloride efflux in response to lubiprostone, which was fully inhibited by GaTx2 and CFTR inhibitor 172 + GlyH-101, respectively. Our study reveals lubiprostone as a pharmacological activator of both CFTR and CLCN2. Overexpression and activation of CLCN2 leads to improved mouse NPD readings, suggesting it is available as an alternative pathway for epithelial chloride secretion in murine airways. The utilization of CLCN2 as an alternative chloride efflux channel could provide clinical benefit to patients with CF, especially if the pharmacological activator is administered as an aerosol.
Eric E, et al. (2013) Dual activation of CFTR and CLCN2 by lubiprostone in murine nasal epithelia. Am J Physiol Lung Cell Mol Physiol. PMID: 23316067
The ClC protein family includes voltage-gated chloride channels and chloride/proton exchangers. In eukaryotes, ClC proteins regulate membrane potential of excitable cells, contribute to epithelial transport, and aid in lysosomal acidification. Although structure/function studies of ClC proteins have been aided greatly by the available crystal structures of a bacterial ClC chloride/proton exchanger, the availability of useful pharmacological tools, such as peptide toxin inhibitors, has lagged far behind that of their cation channel counterparts. Here we report the isolation, from Leiurus quinquestriatus hebraeus venom, of a peptide toxin inhibitor of the ClC-2 chloride channel. This toxin, GaTx2, inhibits ClC-2 channels with a voltage-dependent apparent K(D) of approximately 20 pm, making it the highest affinity inhibitor of any chloride channel. GaTx2 slows ClC-2 activation by increasing the latency to first opening by nearly 8-fold but is unable to inhibit open channels, suggesting that this toxin inhibits channel activation gating. Finally, GaTx2 specifically inhibits ClC-2 channels, showing no inhibitory effect on a battery of other major classes of chloride channels and voltage-gated potassium channels. GaTx2 is the first peptide toxin inhibitor of any ClC protein. The high affinity and specificity displayed by this toxin will make it a very powerful pharmacological tool to probe ClC-2 structure/function.
Thompson CH, et al. (2009) Isolation and characterization of a high affinity peptide inhibitor of ClC-2 chloride channels. J Biol Chem. PMID: 23316067
