Metabolic proof-reading in Plasmodium berghei: essentiality of phosphoglycolate phosphatase

Abstract

12Plasmodium falciparum (Pf) 13 4-nitrophenylphosphatase was earlier shown to be 14 involved in vitamin B1 metabolism by Knöckel et 15 al., (Mol. Biochem. Parasitol. 2008, 157, 241-16 243). An independent BLASTp search showed that 17 the protein had significant homology with phos-18 phoglycolate phosphatase from mouse, human and 19 yeast, and prompted us to re-investigate the bio-20 chemical properties of the recombinant Plasmod-21 ium enzyme. Owing to the insoluble nature of 22 the Pf enzyme, an extended substrate screen and 23 biochemical characterization was performed on its 24 P. berghei (Pb) homolog that led to the identi-25 fication of 2-phosphoglycolate and 2-phospho L-26 lactate as the relevant physiological substrates. 2-27 phosphoglycolate is known to be generated during 28 repair of damaged DNA ends whereas, 2-phospho 29 L-lactate is a product of pyruvate kinase side reac-30 tion. These metabolites are potent inhibitors of the 31key glycolytic enzymes, triosephosphate isomerase 32 and phosphofructokinase, and hence clearance of 33 these toxic metabolites is vital for cell survival and 34 functioning. Gene knockout studies conducted in 35 P. berghei revealed the essential nature of this con-36 served 'metabolic proof-reading enzyme'. 37 Haloacid dehalogenase superfamily 38 (HADSF) is a large family of enzymes consisting 39 mainly of phosphatases and phosphotransferases, 40 that are both intracellular and extracellular in na-41 ture. These enzymes are characterized by the pres-42 ence of a core Rossmanoid-fold and a cap-domain 43 (1, 2). Studies on HADSF members have focused 44 on identifying their physiological substrates by 45 screening a wide range of metabolites that include 46 sugar phosphates, lipid phosphates, nucleotides as 47 well as phosphorylated amino acids and co-factors. 48 This approach has helped understand the physiolog-49 ical relevance of these enzymes in various cellular 50 processes such as cell wall synthesis, catabolic and 51 anabolic pathways, salvage pathways, signaling 52 pathways and detoxification (3-13). Apart from 53 dephosphorylating metabolites, HADSF members 54 have also been know to dephosphorylate proteins 55 and such members are characterized by the absence 56 of the cap domain (1, 2). A large scale study re-57 ported by Huang et al., has identified a HADSF 58 member from Salmonella enterica that catalyzes 59 dephosphorylation of more than 100 phosphory-60 lated substrates (5). This extended substrate speci-61 ficity is a common observation in HADSF members 62 and often leads to a confounding situation where 63 determining the physiological substrate of such 64 promiscuous enzymes becomes a challenging task. 65 Recent studies have identified and char-66 acterised HADSF members from the apicom-67 plexan parasite, Plasmodium (4, 10, 13-16). 68 HADSF members from Plasmodium have been 69 1 P. berghei phosphoglycolate phosphatase found to be involved in processes that lead to 70 the development of resistance to the drug fos-71 midomycin, which inhibits isoprenoid biosynthe-...