Management of the cattle tick, Rhipicephalus microplus, presents a challenge because some populations of this cosmopolitan and economically important ectoparasite are resistant to multiple classes of acaricides. Cytochrome P450 oxidoreductase (CPR) is part of the cytochrome P450 (CYP450) monooxygenases that are involved in metabolic resistance by their ability to detoxify acaricides. Inhibiting CPR, the sole redox partner that transfers electrons to CYP450s, could overcome this type of metabolic resistance. This report represents the biochemical characterisation of a CPR from ticks. Recombinant CPR of R. microplus (RmCPR), minus its N-terminal transmembrane domain, was produced in a bacterial expression system and subjected to biochemical analyses. RmCPR displayed a characteristic dual flavin oxidoreductase spectrum. Incubation with nicotinamide adenine dinucleotide phosphate (NADPH) lead to an increase in absorbance between 500 and 600 nm with a corresponding appearance of a peak absorbance at 340-350 nm indicating functional transfer of electrons between NADPH and the bound flavin cofactors. Using the pseudoredox partner, kinetic parameters for both cytochrome c and NADPH binding were calculated as 26.6 +/- 11.4 mu M and 7.03 +/- 1.8 mu M, respectively. The turnover, K-cat, for RmCPR for cytochrome c was calculated as 0.08 s(-1) which is significantly lower than the CPR homologues of other species. IC50 (Half maximal Inhibitory Concentration) values obtained for the adenosine analogues 2'', 5'' ADP, 2''- AMP, NADP(+)and the reductase inhibitor diphenyliodonium were: 140, 82.2, 24.5, and 75.3 mu M, respectively. Biochemically, RmCPR resembles CPRs of hematophagous arthropods more so than mammalian CPRs. These findings highlight the potential of RmCPR as a target for the rational design of safer and potent acaricides against R. microplus.
[1]Elizade Univ, Dept Biol Sci, Biotechnol Programme, Ilara Mokin, Ondo State, Nigeria.
[2]Northumbria Univ Newcastle, Fac Hlth & Life Sci, Dept Appl Sci, Newcastle Upon Tyne NE1 8ST, England.
[3]USDA, ARS, Knipling Bushland US Livestock Insects Res Lab, Kerrville, TX USA.
[4]USDA, ARS, Vet Pest Genom Ctr, Kerrville, TX USA.
[5]City Univ Hong Kong, Jockey Club Coll Vet Med & Life Sci, Dept Publ Hlth & Infect Dis, Kowloon Tong, Hong Kong, Peoples R China.
[6]Northumbria Univ, Fac Hlth & Life Sci, St Georges Int Sch Med, Dept Biochem & Genet, Newcastle Upon Tyne NE1 8ST, England.
[7]USDA, ARS, San Joaquin Valley Agr Sci Ctr, Parlier, CA USA.
[8]Massey Univ, Sch Vet Sci Genet & Mol Biol, Private Bag 11 222, Palmerston North 4442, New Zealand.
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