Interaction of Cholera Toxin B Subunit with Rat Intestinal Epithelial Cells

We prepared ¹²⁵I-labeled cholera toxin B subunit (¹²⁵I-labeled CT-B, specific activity 98 Ci/mmol) and found that its binding to rat IEC-6 intestinal epithelial cells was high-affinity (K_d 1.9 nM). The binding of labeled protein was completely inhibited by unlabeled thymosin-α₁ (TM-α₁), interferon-α₂ (IFN-α₂), and synthetic peptide LKEKK, which corresponds to residues 16–20 in TM-α₁ and 131–135 in IFN-α₂ (K_i 1.2, 0.9, and 1.6 nM, respectively), but was not inhibited by synthetic peptide KKEKL with inverted amino acid sequence (K_i > 10 μM). Thus, TM-α₁, IFN-α₂, and the LKEKK peptide bind with high affinity and specificity to CT-B receptor on rIEC-6 cells. It was found that CT-B and the LKEKK peptide at concentrations of 10–1000 nM increased nitric oxide production and soluble guanylate cyclase activity in the cells in a dose-dependent manner.