Purification of Plasmodium lophurae cathepsin D and its effects on erythrocyte membrane proteins.
The cathepsin D of Plasmodium lophurae was purified using a combination of CM-Sephadex, pepstatin-agarose and Sephadex G-100 chromatography. The plasmodial enzyme was distinct from that of the host red cell and bovine spleen in its low isoelectric point (pI 4.3). The cathepsin D of P. lophurae, as well as plasmodial extracts demonstrating such proteinase activity, were able to digest the membrane proteins of duckling and human red cells at pH 7.4; proteolysis was not inhibited in phosphate-buffered saline by 100 microM pepstatin. Membrane proteins most susceptible to proteolysis were those of the cytoskeleton, notably bands 1 and 2 (spectrin), bands 2.1-2.6 (spectrin-binding proteins) and band 3. Membrane protein degradation by crude plasmodial extracts was partially inhibited by a combination of 10 mM FeCl3, and 10 mM phenylmethylsulfonyl fluoride in phosphate-buffered saline. The changes induced in erythrocyte membrane proteins by exposure to plasmodial cathepsin D parallel the alterations observed in red cell membranes obtained from malaria infected cells. Since the action of the plasmodial protease was confined to the inner surface of the red cell membrane, it is possible that protease-induced modifications in the red cell cytoskeleton could lead to merozoite release.
D000818:Animals; D001798:Blood Proteins / Q000378:metabolism; D002402:Cathepsin D; D002403:Cathepsins / Q000302:isolation & purification* / Q000494:pharmacology; D004372:Ducks; D004591:Electrophoresis, Polyacrylamide Gel; D004910:Erythrocyte Membrane / Q000187:drug effects*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000097:blood*; D010436:Pepstatins / Q000494:pharmacology; D010961:Plasmodium / Q000201:enzymology*; D013154:Spleen / Q000201:enzymology
D001798:Blood Proteins; D008565:Membrane Proteins; D010436:Pepstatins; C014195:Streptomyces pepsin inhibitor; D002403:Cathepsins; D002402:Cathepsin D; C031375:pepstatin