Pyruvate kinase type-II isozyme in Plasmodium falciparum localizes to the apicoplast

Abstract

Bioinformatics research on Plasmodium falciparum revealed two isoforms of pyruvate kinase: type-I and type-II enzymes. The type-I enzyme shows typical glycolytic properties, while type-II enzyme is involved in fatty acid type-II biosynthesis and has been predicted to localize to the apicoplast with the targeting signal in its N-terminus. The type-I and type-II isoforms have the same evolutionary origin as Toxoplasma gondii isozymes, TgPyKI and TgPyKII, respectively; however, TgPyKII localizes to both the mitochondrion and the apicoplast. Accordingly, we made a recombinant full length of P. falciparum pyruvate kinase type-II protein using a wheat germ cell-free expression system and obtained a specific antibody against the type-II protein. Fluorescent microscopic analysis revealed that P. falciparum type-II enzyme was localized only to the apicoplast, not to the mitochondrion. The data suggest differences in localization and metabolic pathways between P. falciparum and T. gondii pyruvate kinase isoforms.

Stage-Specific Changes in Plasmodium Metabolism Required for Differentiation and Adaptation to Different Host and Vector Environments

“…PEPCK-derived PEP can be converted to pyruvate in the cytosol by the glycolytic enzyme, pyruvate kinase 1 [58] and be further catabolized in the mitochondrion TCA cycle [13] generating the ATP required for multiple rounds of DNA replication in sporogony. Alternatively it can enter the apicoplast where it can be converted to pyruvate and acetyl-CoA by pyruvate kinase 2 [59] and apicoplast-resident pyruvate dehydrogenase [60], respectively (Fig 2A). Apicoplast-derived acetyl-CoA is required for de novo fatty acid synthesis during sporozoite development in the mosquito by the FASII pathway in P .…”

Fatty acid metabolism in the Plasmodium apicoplast: Drugs, doubts and knockouts

“…Like the pPTs, PKII is expected to support multiple aspects of apicoplast metabolism, providing pyruvate for both the FASII and DOXP pathways and ATP for a range of enzymatic reactions [112,117]. The apicoplast localization of the P. falciparum and T. gondii PKII have been confirmed by antibody labeling and epitope tagging [115,120], and the activity of T. gondii enzyme has been demonstrated in vitro [62]. Interestingly, the TgPKII displayed an exclusive preference for GDP over ADP as a phosphate acceptor [62], distinguishing it from most other pyruvate kinases.…”

“…Pyruvate kinases catalyze the transfer of a phosphate group from PEP to ADP, forming pyruvate and ATP. Plasmodium and T. gondii possess two pyruvate kinases, a type I isoform (PKI) involved in glycolysis in the cytoplasm, and the type II isoform (PKII) of the apicoplast [115,119,120]. Like the pPTs, PKII is expected to support multiple aspects of apicoplast metabolism, providing pyruvate for both the FASII and DOXP pathways and ATP for a range of enzymatic reactions [112,117].…”

An overview of structure, function, and regulation of pyruvate kinases

“…Cluster I PyKs are regulated by FBP or FDP while AMP or sugar monophosphates modulate the enzymes in cluster II . PyKs belonging to both clusters are expressed in gamma‐proteobacteria, and in apicomplexan parasites in which cluster I and cluster II PyKs are found in cytosolic and apicoplast compartments, respectively . Organisms carrying both families of PyKs include a number of clinically significant pathogens such as Escherichia coli , Vibrio cholera , Salmonella typhimurium , Toxoplasma gondii , and Plasmodium falciparum …”

“…PyKII of T. gondii is also localized in the mitochondria in addition to the apicoplast. PKI is similar to the glycolytic form of PyKs whereas PyKII is implicated in fatty acid biosynthesis . Notably, PyKII sequences possess a Lys residue in the position corresponding to Glu117 in rabbit muscle PyK, and therefore group with monovalent cation‐independent enzymes.…”