This chapter elaborates the structural chemistry and the biological aspects of plasmepsins. Plasmepsin I is capable of cleaving native human hemoglobin and acid-denatured globin, with a pH optimum around 5. Plasmepsin I, II, IV and HAP are made as 51 kDa precursors that are cleaved to 37 kDa mature forms. There are two disulfide bonds predicted. The second domain contains an active-site Asp-Ser-Gly instead of the usual aspartic protease Asp-Thr-Gly. HAP is an exception to this. A molecular model of plasmepsin I has been constructed, based on the crystal structure of the highly homologous plasmepsin II. The modeled structure bears general features of mammalian and fungal aspartic proteases, and among mammalian orthologs shares greatest structural similarity with cathepsin D. Pepstatin is bound in the active site with enough conformational difference from mammalian homologs to give hope for development of a selective inhibitor. Native plasmepsin I, II and HAP can be prepared in nanogram to low microgram amounts from cultured intraerythrocytic organisms. Plasmepsin I, II, IV and HAP are 55–75% identical at the amino acid level.