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Expression, Characterization, and Cellular Localization of Knowpains, Papain-Like Cysteine Proteases of the Plasmodium knowlesi Malaria Parasite

Figure 2

Production of recombinant knowpains.

Knowpains were expressed in BL21(DE3) E. coli cells, purified from induced cells under denaturing conditions, refolded, and processed into mature proteases. For indicated knowpains (knowpain-2, KP2; knowpain-3, KP3; and knowpain-4, KP4) samples of the total lysates of uninduced (Un) and induced (In) cells, soluble (Sol) and insoluble (Ins) fractions of induced cells, Ni-NTA purified (Ni) and ion-exchange purified (IE) proteins, and refolded (Ref) and processed mature proteases (Pros) were resolved by 12% SDS-PAGE under reducing conditions. The gel was stained with coomassie blue. The same marker (M) was used in all cases, and molecular masses of markers are indicated in kDa; and the position of 29 kDa marker for the processed KP3 lane is indicated.

Figure 2