Abstract 1. 1. Essential differences in the binding of NADP-dependent malic enzymes from mammals, fish and crustacea to 2′,5′ADP-Sepharose 4B have been shown. 2. 2. The enzymes from mammalian and fish tissues interact with 2′,5′ADP-Sepharose, whereas the enzymes from crayfish Orconectes limosus and shrimp Crangon crangon tissues do not. 3. 3. These results provide evidence that 2′,5′ADP-Sepharose cannot be used for the purification of NADP-dependent malic enzyme from crayfish and shrimp and indicate presumably a structural difference in the NADP-binding site of crustacean enzyme.