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Non-canonical maturation of two papain-family proteases in Toxoplasma gondii.

The Journal of biological chemistry (2012-12-20)
Zhicheng Dou, Isabelle Coppens, Vern B Carruthers
ABSTRACT

Proteases regulate key events during infection by the pervasive intracellular parasite Toxoplasma gondii. Understanding how parasite proteases mature from an inactive zymogen to an active enzyme is expected to inform new strategies for blocking their actions. Herein, we show that T. gondii cathepsin B protease (TgCPB) does not undergo self-maturation but instead requires the expression of a second papain-family cathepsin protease, TgCPL. Using recombinant enzymes we also show that TgCPL is capable of partially maturing TgCPB in vitro. Consistent with this interrelationship, antibodies with validated specificity detected TgCPB in the lysosome-like vacuolar compartment along with TgCPL. Our findings also establish that TgCPB does not localize to the rhoptries as previously reported. Accordingly, rhoptry morphology and rhoptry protein maturation are normal in TgCPB knock-out parasites. Finally, we show that although maturation of TgCPL is independent of TgCPB, it may involve an additional protease(s) in conjunction with self-maturation.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Papain from papaya latex, lyophilized powder, aseptically filled
Sigma-Aldrich
Cathepsin B from bovine spleen, lyophilized powder, ≥10 units/mg protein
Sigma-Aldrich
Cathepsin B from human liver, buffered aqueous solution, ≥1,500 units/mg protein (E1%/280)
Sigma-Aldrich
Papain from papaya latex, crude powder, 1.5-10 units/mg solid
Sigma-Aldrich
Papain from Carica papaya, solution, light brown, ≥10 U/mg protein (~25 mg/ml)
Sigma-Aldrich
Papain from Carica papaya, powder, ≥3 U/mg
Sigma-Aldrich
Cathepsin B from human placenta, lyophilized powder, ≥5 units/mg protein
Sigma-Aldrich
Cathepsin L from human liver, ≥0.5 units/mg protein, solution