Structure and substrate fingerprint of aminopeptidase P from Plasmodium falciparum

Abstract

Malaria is one of the world's most prevalent parasitic diseases, with over 200 million cases annually. Alarmingly, the spread of drug-resistant parasites threatens the effectiveness of current antimalarials and has made the development of novel therapeutic strategies a global health priority. Malaria parasites have a complicated lifecycle, involving an asymptomatic 'liver stage' and a symptomatic 'blood stage'. During the blood stage, the parasites utilise a proteolytic cascade to digest host hemoglobin, which produces free amino acids absolutely necessary for parasite growth and reproduction. The enzymes required for hemoglobin digestion are therefore attractive therapeutic targets. The final step of the cascade is catalyzed by several metalloaminopeptidases, including aminopeptidase P (APP). We developed a novel platform to examine the substrate fingerprint of APP from Plasmodium falciparum (PfAPP) and to show that it can catalyze the removal of any residue immediately prior to a proline. Further, we have determined the crystal structure of PfAPP and present the first examination of the 3D structure of this essential malarial enzyme. Together, these analyses provide insights into potential mechanisms of inhibition that could be used to develop novel antimalarial therapeutics.

P1′ Residue-Oriented Virtual Screening for Potent and Selective Phosphinic (Dehydro) Dipeptide Inhibitors of Metallo-Aminopeptidases

“…The M1 family (Table 3) includes, for example, the enzyme from Plasmodium falciparum, the most fatal of the four protozoan species that cause malaria in humans. This species also produces an M17 aminopeptidase [93], an aspartyl aminopeptidase classified in the M18 family [36], and the M24 aminopeptidase P [51]. Metallo-aminopeptidases are etiological factors of microbial infections, for example, meningitis caused by Neisseria meningitides, or yeast species.…”

“…M17 mixed type A/I aminopeptidase from Helicobacter pylori, in addition to urease (Nobel Prize in Medicine for Marshall and Warren in 2005 [95]), is among the potential targets for organophosphorus inhibitors. This bizinc-dependent enzyme has a wide active center arranged for peptides with long side chains [51], but the S1 preferences have not yet been specified. Nevertheless, good docking results were obtained for compounds with boronyl groups ( Table 7).…”

Structure–Function Relationship of Aminopeptidase P from Pseudomonas aeruginosa

“…Several APPro structures have been reported to date, including APPro from Escherichia coli (Ec-PepP, 1M35) ( Wilce et al, 1998 ), Streptococcus thermophilus (3IL0), Bacillus anthracis (3IG4), Yersinia pestis (4PV4), Thermotoga maritima (2ZSG), Streptococcus pyogenes (3OVK), human (hAPP1, 3CTZ) ( Li et al, 2008 ), Caenorhabditis elegans (CeAPP1, 4S2R) ( Iyer et al, 2015 ) and Plasmodium falciparum (PfAPP, 5JQK) ( Drinkwater et al, 2016 ). Ec-PepP shares the highest sequence identity with Pa-PepP and both of them are prokaryotic APPro ( Gonzales and Robert-Baudouy, 1996 ).…”

“…In contrast to the prolidases that cleave dipeptides only, APPro is capable of modifying protein substrates. The relatively larger substrate binding ability of APPro is partly due to the extension of the substrate binding site beyond the S1’–S1 region ( Drinkwater et al, 2016 ). The modeled tripeptide-bound Pa-PepP structure indicated that Arg153 and Arg351 accommodate the carboxylate group of the P2’ Leu, and guide the C-terminus of the substrate toward the tetramer surface ( Figure 5A ).…”

Structures and activities of widely conserved small prokaryotic aminopeptidases‐P clarify classification of M24B peptidases

“…[10][11][12] Aminopeptidase-P from Plasmodium falciparum is an essential enzyme and is proposed to have a role in vacuolar catabolism of hemoglobin and cytosolic peptide turnover, and it has been identified as a potential drug target. 13,14 In bacteria, aminopeptidases-P have been shown to be important for the recycling of proline in Salmonella typhimurium. 15 In Bacillus subtillis, the genes encoding aminopeptidase-P (pepA) and prolidase (pepB) have been shown to be involved in increasing the concentration of Lproline inside the cells leading to their osmoprotection.…”