m1-Toxin isotoxins from the green mamba (Dendroaspis angusticeps) that selectively block m1 muscarinic receptors

The venom of the green mamba, Dendroaspis angusticeps, was found to have at least six trace m1-specific toxins that block the binding of H-3-N-methylscopolamine to cloned mi muscarinic receptors. Four were isolated by gel filtration, cation exchange HPLC and reversed-phase HPLC and named m1-toxins1-4. Recovery:was 180, 90, 20 and 10 mu g/g dry venom, respectively. m1-Toxin1 (the original ml-toxin) was found to have the sequence, L T C V K S N S I W F P T S E D C P D G Q N L C F R W Q Y I S P R M Y D F T R G C A A T C P K A E Y R D V I N C C G T D K C N K, calculated mass = 7473 Da and calculated pI = 8.2. This sequence had been predicted previously from a cDNA cloned from the venom glands of this snake. The binding of m1-toxin1 was irreversible, so its K-d could not be determined, m1-Toxin2 differed only in proline-19, mass = 7455 and pI = 8.5. Partial sequence data for m1-toxin3 showed aspartate-7 and m1-toxin4 showed isoleucine-12, asparagine-16 and alanine-19. m1-Toxins1-4 have seven conserved amino acids not found in homologous mamba toxins that bind to other muscarinic receptors (MT1, MT2, m4-toxin = MT3, MT4, MT5, MT alpha and MT beta). Some of these residues may be essential for m1-specificity. Since m1-toxin1 binds irreversibly in artificial cerebrospinal fluid at 37 degrees C, it is a particularly attractive antagonist for studies in vivo. (C) 1999 Elsevier Science Ltd. All rights reserved.