Falkenstein_1999_J.Protein.Chem_18_233

Fasciculins (Fas) are three-looped polypeptides isolated from mamba venom which exert their toxic action by inhibiting noncompetitively acetylcholinesterase (AChE). A peptide (Fas-D) encompassing the first loop sequence was synthesized and characterized chemically, structurally, and functionally. Fas-D possesses an intramolecular disulfide bridge, present in the native toxin. Circular dichroism (CD) indicated the existence of 21.8% beta-sheet content and 24.2% beta-turn in this peptide, compatible with crystallographic data of the native toxin. The peptide showed only low partial AChE inhibition at submillimolar concentrations, much lower than that observed with Fas and a peptide (Fas-B) encompassing the second loop sequence. The simultaneous presence of Fas-D and Fas-B produced an additive inhibitory effect on AChE activity; calculated Ki and alphaKi values (7.3 +/-2.4 microM and 10.0 +/- 1.8 microM, respectively) were not significantly different, thus indicating noncompetitive inhibition. These results are consistent with site-directed mutagenesis studies and analysis of the crystal structure of the Fas-AChE complex, which indicate that residues from loops I and II contribute to Fas binding to the enzyme.