From a Phoneutria nigriventer venom gland cDNA library several clones coding for the insect specific neurotoxin Tx4(6-1) were isolated. cDNA analysis showed that the encoded protein contained three distinct segments, comprising a signal sequence of 16 amino acids, followed by a glutamate-rich sequence of 18 amino acids and, finally, the coding region for the mature toxin. The deduced amino acid sequence for the mature polypeptide was identical to the protein sequence determined chemically. In addition, two new putative toxins called Pn4A and Pn4B were characterized and their predicted complete amino acid sequence revealed approximately 78% similarity to Tx4(6-1).