Glutathione S-transferase of
Plasmodium falciparum (PfGST) displays a peculiar dimer to
tetramer transition that causes full
enzyme inactivation and loss of its ability to sequester parasitotoxic
hemin. Furthermore, binding of
hemin is modulated by a cooperative mechanism.
Site-directed mutagenesis, steady-state kinetic experiments, and
fluorescence anisotropy have been used to verify the possible involvement of loop 113-119 in the tetramerization process and in the cooperative phenomenon. This protein segment is one of the most prominent structural differences between PfGST and other
GST isoenzymes. Our results demonstrate that truncation, increased
rigidity, or even a simple
point mutation of this loop causes a dramatic change in the tetramerization kinetics that becomes at least 100 times slower than in the native
enzyme. All of the
mutants tested have lost the positive
cooperativity for
hemin binding, suggesting that the integrity of this peculiar loop is essential for intersubunit communication. Interestingly, the tetramerization process of the native
enzyme that occurs rapidly when
GSH is removed is prevented not only by
GSH but even by oxidized
glutathione. This result suggests that protection by PfGST against
hemin is independent of the redox status of the
parasite cell. Because of the importance of this unique segment in the function/structure of PfGST, it could be a new target for the development of
antimalarial drugs.
