Mapping Receptor Sites for Sodium Channel Blocking Insecticides DCJW and Metaflumizone in an Insect Sodium Channel

Abstract

Acquired pest resistance to long-used insecticides, such as sodium channel agonists DDT and pyrethroids, motivates development of new insecticides with different mechanisms of action. Indoxacarb, its insect-specific active metabolite DCJW, and metaflumizone (MFZ) are sodium channel blocker insecticides (SCBIs). They preferably bind to and trap sodium channels in the slow-inactivated state, a mode of action similar to that of local anesthetics (LAs). Previous studies identified several SCBI-sensing residues that face the inner pore, but the atomic mechanism of action of SCBIs is unknown. Unlike classical LAs, DCJW and MFZ are uncharged molecules. They may chelate a permeant ion to form a cationic blocking particle. Here we have built a homology model of the open cockroach sodium channel BgNav1-1 using as templates the X-ray structures of an open potassium channel Kv1.2 and a closed sodium channel NavAb. We then generated libraries of conformers of sodium ion-bound SCBIs and docked them into the channel model imposing ligand-sidechain distance constraints between SCBIs and known SCBI-sensing residues. Our computations predicted that SCBIs bind in the inner pore with the SCBI-chelated sodium ion located at the cation-attractive focus of P1 helices and an aromatic moiety extended into the III/IV repeat interface that has insect-specific residues. Model-driven mutagenesis and electrophysiology identified five new SCBI-sensing residues, including residues that are specific to either DCJW or MFZ. Our study supports the following conclusions. (1) Electroneutral SCBIs chelate a sodium ion by the common C=N-NH-C=O fragment to form a cationic blocking particle. (2) The blocking particle preferably binds in the inner pore of sodium-deficient slow-inactivated channels. (3) Selective toxicity of MFZ is due to binding to insect-specific residues in the III/IV repeat interface. Supported by NIH and NSERC.