AbstractThe 33,000 Dalton venom protein ofChelonusnearcurvimaculatuswas characterized for structural properties of charge, quaternary associations, and relationship to polydnavirus encoded proteins. Homogenous isoforms of the protein were isolated from the venom by sequential steps of (1) microdissection, (2) separation based on charge (Mono‐Q column HPLC or narrow‐range electrofocusing), and (3) centrifugal filtration based on molecular weight using Centricon microconcentrators. The purified protein dimerized under native conditions, and this quaternary association became denaturation resistant under certain conditions. Chemical modification of lysine epsilon amino groups did not disrupt such dimerization. The cDNA for the protein did not possess high similarity to any sequence encoded in the polydnavirus, as indicated by results of Southern blotting, but does possess similarity in its repeats to the repeats of the immunologically protective surface glycoprotein ofLeishmania amazonensis. © 1994 Wiley‐Lis