Thioredoxin reductase and dihydrolipoamide dehydrogenases of Plasmodium falciparum

Paul James McMillan
2006
Plasmodium falciparum is an obligate intracellular protozoan parasite and is the causative agent of malaria, which infects 270 million people and causes 2-3 million human deaths each year. During the erythrocytic stages of their life cycle the parasites are exposed to reactive oxygen species generated by the host and their own metabolism. As a result the parasites have developed efficient antioxidant defence mechanisms, which include the thioredoxin system. A key component of the thioredoxin
more » ... tem is the flavin adenine dinucleotide (FAD)-dependent disulphide oxidoreductase thioredoxin reductase (PfTrxR), which provides the reducing power for the system using NADPH. PfTrxR is a high Mr TrxR and has been genetically and chemically validated as a potential drug target by targeted gene dismption and the identification of several specific inhibitors. PfTrxR contains three redox active centres (FAD, Cys88/Cys93 and Cys535/Cys540) that are in redox communication. This study examined the catalytic mechanism (i.e. the transfer of reducing equivalents between these redox active centres) of PfTrxR by employing pre-steady state kinetics of the reductive (reduction of PfTrxR by NADPH) and oxidative half reaction (oxidation of reduced PfTrxR by thioredoxin). This identified that the PfTrxR protein conformed to the catalytic mechanism for high Mr TrxR, which was proposed from studies on the Drosophila melanogaster TrxR (DmTrxR). However, a few key differences were observed between the proteins from the two organisms. Firstly, the reductive half reaction of PfTrxR was observed to proceed at a faster rate than that of DmTrxR. Secondly, the oxidative half reaction was observed to proceed at a slower rate than that of DmTrxR protein. Finally, PfTrxR was shown not to cycle between a two (EH2) and four (EH4) electron reduced species in vitro (as demonstrated in DmTrxR) and was observed to be almost fully oxidised following reaction with excess thioredoxin. However, the rate of oxidation from the two electron reduced (EH2) to oxidise [...]
doi:10.5525/gla.thesis.71027 fatcat:l5zrzezzh5c2vjq437jb6u7cyi