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Figure 5 | Malaria Journal

Figure 5

From: Characterisation and expression of a PP1 serine/threonine protein phosphatase (PfPP1) from the malaria parasite, Plasmodium falciparum: demonstration of its essential role using RNA interference

Figure 5

Constitutive expression of parasitic PfPP1. (A) Western blot: Total protein (80 μg) from the ring (R), Trophozoite (T), schizont (S), and early (G1) and late (G2) sexual stages of Pf were probed with a mixture of anti-PP1 and anti-Pfg27 antibodies as described [14]. (B) Microcystin-sepharose chromatography: About 500 μg of the following extracts was subjected to microcystin affinity chromatography and the bound proteins analyzed by Western blot using PP1 antibody: extract of uninfected RBC processed identically (lane 1); Pf extract (lane 2); Pf extract pre-incubated with 1 μM microcystin-LR at room temperature for 5 min (lane 3); unbound fraction (a double-pass flow-through from the column) (lane 4). Recombinant His-tagged PfPP1 is displayed in lane 5 for comparison. The native and recombinant PP1 bands are marked by open and closed arrows, respectively. Sizes of protein standards are indicated on the left. Two non-PP1 proteins are also seen in the blot in panel B. The ~25 kDa band (common in lanes 1, 2, and 3) is evidently a RBC protein. The ~40 kDa band (lanes 2, 3), on the other hand, is a Plasmodium protein, since it is absent in the RBC fraction. We speculate that these proteins non-specifically bound to the Sepharose matrix, since they could not be competed out by microcystin (lane 3).

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