Three aspartic proteinases with similar Mr values (approx. 80,000) but from distinct sources (human gastric mucosa, human erythrocyte membranes and rat spleen) were shown to have immunological cross-reactivity and comparable mobilities when subjected to polyacrylamide-gel electrophoresis under non-denaturing conditions. Kinetic parameters (kcat, Km and Ki) were determined for the interactions of the three enzymes with two synthetic chromogenic substrates and five inhibitors (naturally occurring and synthetic). On this basis it would appear that all of the enzymes should be considered equivalent to cathepsin E. pH-activity measurements indicated that the aspartic proteinase that originated from the erythrocyte membranes retained activity at a higher pH value than either of its readily soluble counterparts.