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Biotechnology and Applied Biochemistry (2002) 35, (9–17) (Printed in Great Britain)

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Resolving the DDT target protein in insects as a subunit of the ATP synthase

Hassan M. Younis¹, Mahmoud M. Abo-El-Saad, Reda K. Abdel-Razik and Samia A. Abo-Seda

Laboratory of Bioenergetics and Biochemical Toxicology, Department of Pesticide Chemistry, Faculty of Agriculture, El-Chatby, 21545, Alexandria University, Alexandria, Egypt

Key words: DDT sensitivity, mode of action, selective insecticide target, structure-mediated resistance mechanism.

Abbreviations used: DDT, 1,1-bis-(p-chlorophenyl)-2,2,2-trichloroethane; DDE, 1,1-bis-(p-chlorophenyl)-2,2-dichloroethylene; DDA, bis-(p-chlorophenyl)acetic acid; methoxychlor, 1,1-bis-(p-methoxyphenyl)-2,2,2-trichloroethane; DCCD, N, N-dicyclohexylcarbodi-imide; PEP, phosphoenolpyruvate; PK, pyruvate kinase.

¹To whom correspondence should be addressed (e-mail hmyounis@dataxprs.com.eg).

1,1-bis-(p-Chlorophenyl)-2,2,2-trichloroethane (DDT) inhibited the ATP hydrolytic activity of the ATP synthase from a DDT-susceptible insect (Apis mellifera) as well as a DDT-tolerant insect (Spodoptera littoralis), and from rat liver and bovine heart in a parallel way to its insecticidal properties and selectivity of action. Inhibition of the ATPase activity of these preparations by DDT was parallel to the poisoning of the source organism with DDT. Furthermore, both the inhibition and poisoning of insects were affected similarly by temperature. Inhibition of the insect enzyme activity by DDT was specific and differed from that by oligomycin or N,N-dicyclohexylcarbodi-imide (DCCD). PAGE analysis of the various preparations of the enzyme showed that the inhibition of the enzyme activity by DDT was associated with the presence of a selective protein band with an apparent molecular mass of 23 kDa. This protein band exists in the preparations from the DDT-susceptible insects but was absent from the preparations of the enzyme from the DDT-insensitive sources. Removal of this protein band from the enzyme rendered its activity insensitive to inhibition by DDT. The protein was purified directly from mitochondria and the DDT sensitivity was reconstituted upon its addition to the DDT-insensitive F₁-ATPase. We conclude that this identified protein of the ATP synthase is the DDT target protein in insects.

Received 23 July 2001/21 September 2001; accepted 12 October 2001

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