IDENTIFICATION OF A CALMODULIN‐BINDING SITE WITHIN THE DOMAIN I OF B ACILLUS THURINGIENSIS C ry3 A a TOXIN

B acillus thuringiensis C ry3 A a toxin is a coleopteran specific toxin highly active against C olorado P otato B eetle ( CPB). We have recently shown that C ry3 A a toxin is proteolytically cleaved by CPB midgut membrane associated metalloproteases and that this cleavage is inhibited by ADAM metalloprotease inhibitors. In the present study, we investigated whether the C ry3 A a toxin is a calmodulin ( C a M ) binding protein, as it is the case of several different ADAM shedding substrates. In pull‐down assays using agarose beads conjugated with C a M , we demonstrated that C ry3 A a toxin specifically binds to C a M in a calcium‐independent manner. Furthermore, we used gel shift assays and 1 H NMR spectra to demonstrate that C a M binds to a 16‐amino acid synthetic peptide corresponding to residues N 256‐ V 271 within the domain I of C ry3 A a toxin. Finally, to investigate whether C a M has any effect on C ry3 A a toxin CPB midgut membrane associated proteolysis, cleavage assays were performed in the presence of the C a M ‐specific inhibitor trifluoperazine. We showed that trifluoperazine significantly increased C ry3 A a toxin proteolysis and also decreased C ry3 A a larval toxicity.