MOLECULAR-CLONING, EXPRESSION AND ENZYMATIC-ACTIVITY OF A THIOREDOXINPEROXIDASE FROM DIROFILARIA-IMMITIS

A Dirofilaria immitis cDNA clone encoding a nucleic acid homolog of th ioredoxin peroxidase (nDiTPx) was isolated from a fourth-stage larval cDNA library, using serum from dogs vaccinated by chemotherapeutically -abbreviated D., immitis larval infections. The protein encoded by nDi TPx had a predicted molecular mass of 22.1 kDa and the deduced amino a cid sequence was homologous to thioredoxin peroxidase-like sequences d escribed in other filarial nematodes, yeast, bacteria and mammals. As is true for other members of this peroxiredoxin family, the nDiTPx-enc oded protein had the conserved cysteine near the amino terminus, consi dered to be essential for enzyme activity. nDiTPx was expressed in E. coli and the resulting recombinant fusion protein was shown to have th ioredoxin peroxidase (TPx) activity, by its ability to protect DNA fro m oxidative-nicking in a metal-catalyzed oxidation system. A polyclona l antibody to the DiTPx fusion protein detected a 22-kDa native protei n in D. immitis larval and adult parasite extracts. (C) 1997 Elsevier Science B.V.