Modulation ofAnopheles gambiaeEpsilon glutathione transferase activity by plant natural productsin vitro

Abstract

Elevated glutathione transferase (GST) E2 activity is associated with DDT resistance in the mosquito Anopheles gambiae. The search for chemomodulators that inhibit the function of AgGSTE2 would enhance the insecticidal activity of DDT. Therefore, we examined the interaction of novel natural plant products with heterologously expressed An. gambiae GSTE 2 in vitro. Five of the ten compounds, epiphyllocoumarin (Tral-1), knipholone anthrone, isofuranonaphthoquinones (Mr 13/2, Mr13/4) and the polyprenylated benzophenone (GG1) were shown to be potent inhibitors of AgGSTE2 with IC 50 values of 1.5 mM, 3.5 mM, 4 mM, 4.3 mM and 4.8 mM respectively. Non-competitive inhibition was obtained for Tral 1 and GG1 with regards to GSH (K i of 0.24 mM and 0.14 mM respectively). Competitive inhibition for Tral1 was obtained with CDNB (K i ¼ 0.4 mM) whilst GG1 produced mixed type of inhibition. The K i and K i ' for GSH for Tral-1 and GG1 were 0.2 mM and 0.1 mM respectively. These results suggest that the novel natural plant products, particularly Tral-1, represent potent AgGSTE2 in vitro inhibitors.

Inactivation ofAnopheles gambiaeGlutathione Transferaseε2 by Epiphyllocoumarin

“…The specific activity of AgGST ε 2 is also comparable with the activities described for other recombinant AgGST ε 2 of the same class, for example, the specific activity of AgGST ε 1 with CDNB as a substrate was reported to be 56.4  μ mol min −1  mg −1 [19]. The molecular weight of AgGST ε 2 obtained was 25 kDa (Figure 2), a value which is the same as the literature value [26]. …”

“…Epoxy activated Sepharose 6B (Pharmacia Biotech, Uppsala, Sweden) affinity gel was used for affinity chromatography purification of AgGST ε 2 as described before [26]. In order to determine the tubes in which most of the enzyme was located, a 1-chloro-2,4-dinitrobenzene (CDNB) assay was carried out.…”

“…The various fractions collected during purification, that is, the cell supernatant, nonbound, affinity pool, and dialysate fractions, were tested for GST activity [26]. The 1 mL reaction mixture consisted of 1 mM glutathione (GSH), 1 mM CDNB, 1 mM EDTA, and 0.1 M sodium phosphate buffer at pH 6.5.…”

“…This leads to spasms and eventual death. Previously, reports suggested that Tral 1 reversibly inhibited GST ε 2 in vitro [26] and so our aim was to determine if Tral 1 can also irreversibly inhibit GST ε 2, resulting in time-dependent effects. These effects would be useful in controlling insecticide resistance in malarial mosquitoes that overexpress this enzyme.…”

Glutathione transferase fromPlasmodium falciparum– Interaction with malagashanine and selected plant natural products

“…It has been reported that GSTs are able to interact covalently and noncovalently with various compounds that are not substrates for enzymatic activity [28]. GST activity has been shown to be modulated by natural plant products [29,30]. Flavonoids have been shown to inhibit GSTs in human blood platelets as well as in cancer cell lines [31].…”