BACKGROUND: Plasmodium falciparum, the causative agent of human malaria, 
expresses two aminopeptidases, PfM1AAP and PfM17LAP, critical to generating a 
free amino acid pool used by the intraerythrocytic stage of the parasite for 
proteins synthesis, growth and development. These exopeptidases are potential 
targets for the development of a new class of anti-malaria drugs.
METHODOLOGY/PRINCIPAL FINDINGS: To define the substrate specificity of 
recombinant forms of these two malaria aminopeptidases we used a new library 
consisting of 61 fluorogenic substrates derived both from natural and unnatural 
amino acids. We obtained a detailed substrate fingerprint for recombinant forms 
of the enzymes revealing that PfM1AAP exhibits a very broad substrate tolerance, 
capable of efficiently hydrolyzing neutral and basic amino acids, while PfM17LAP 
has narrower substrate specificity and preferentially cleaves bulky, hydrophobic 
amino acids. The substrate library was also exploited to profile the activity of 
the native aminopeptidases in soluble cell lysates of P. falciparum malaria.
CONCLUSIONS/SIGNIFICANCE: This data showed that PfM1AAP and PfM17LAP are 
responsible for majority of the aminopeptidase activity in these extracts. These 
studies provide specific substrate and mechanistic information important for 
understanding the function of these aminopeptidases and could be exploited in 
the design of new inhibitors to specifically target these for anti-malaria 
treatment.