The activity of an acid proteinase from mouse red cells infected with Plasmodium berghei is 5 to 10 times that of normal red cells. The difference is 2to 3-fold in rats. Levels of acid phosphatase were similar in infected and uninfected cells. Maximal proteinase activity in vitro requires either repeated freeze-thawing of samples or inclusion of a detergent in the reaction mixture, and the enzyme from infected cells is solubilized much less readily than that from normal cells. Acid phosphatase is readily released from both normal and infected cells. Much of the proteinase is found in parasites freed from host cells by hemolysis. When acid-denatured hemoglobin is used as substrate, the pH optimum from various fractions of both normal and infected cells is 2.5 to 3.0, and little or no activity is observed above pH 4.5. The partially purified enzyme is not affected by high concentrations of either dithiothreitol or EDTA. High concentrations of several antimalarial drugs did not inhibit the enzyme from normal or infected cells. There is considerable evidence to suggest that the malarial parasite obtains the bulk of its amino acids from breakdown of host-cell hemoglobin (McKee, 1951; Fulton and Grant, 1956; Polet et al., 1969; Cenedella et al., 1968). Yet there have been relatively few studies on the enzymes involved in this process. Moulder and Evans (1946) found that cell-free extracts of Plasmodium gallinaceum from chicken red blood cells degraded hemoglobin at a very slow rate when tested at pH 6.5. Denatured globin was hydrolyzed much faster. Cook et al. (1961), studying P. berghei and P. knowlesi, suggested the existence of two proteinases in each species, with pH optima of 4 and 8 (P. berghei) and 5 and 8 (P. knowlesi). In each case, the alkaline protease was the more active and the acid protease was quite unstable. However, the demonstration that parasites prepared by the methods used in that study are contaminated with host cell materials (Cook et al., 1969; Ladda, 1969) raises the possibility that the enzymes studied were not completely of parasite origin, especially in view of the finding that erythrocyte proteases may be tightly membrane-bound (Morrison and Neurath, 1953). In addition, it is not possible, from the data given, to determine if leukocytes were removed prior to disruption of red cells. We have recently begun to reinvestigate some of the properties of proteases of erythroReceived for publication 12 April 1973. cytic stages of Plasmodium berghei. We feel that because of the probable importance of these enzymes, not only in the provision of the cell with nutrients, but also in the structural reorganization that occurs at the onset of the erythrocytic stages, it is important to identify and characterize parasite-specific proteases. Furthermore, it is becoming clear that a given cell may contain a variety of proteinases, which differ in specificity and localization (cf. Barrett and Dingle, 1971). A study of proteinases in Plasmodium may be of particular value since chloroquine inhibits certain proteolytic enzymes (Cowey and Whitehouse, 1966; Woessner, 1969) and accumulates within lysosomes in certain tissues (Allison and Young, 1964; Polet, 1970). It has been uggested that this compound may accumulate i lysosomes in Plasmodium (Polet, 1970; Homewood et al., 1972). Also, autoradiography at the ultrastructural level suggests that primaquine may accumulate in lysosomelike bodies (Aikawa and Beaudoin, 1970). In this paper we report the presence of elevated levels of an acid proteinase in red cells from rats and mice infected with P. berghei. Our data suggest that this enzyme, rather than a neutral or alkaline protease, is the main proteolytic enzyme in the parasite. The enzyme is tightly bound, has cathepsinD-like properties, and is not inhibited by a number of antimalarial drugs. Our data differ in a number of other ways from that of Cook etal. (1961).