title: Adenylosuccinate synthetase and adenylosuccinate lyase deficiencies trigger growth and infectivity deficits in Leishmania donovani.
abstract: Leishmania are auxotrophic for purines, and consequently purine acquisition from the host is a requisite nutritional function for the parasite. Both adenylosuccinate synthetase (ADSS) and adenylosuccinate lyase (ASL) have been identified as vital components of purine salvage in Leishmania donovani, and therefore Δadss and Δasl null mutants were constructed to test this hypothesis. Unlike wild type L. donovani, Δadss and Δasl parasites in culture exhibited a profoundly restricted growth phenotype in which the only permissive growth conditions were a 6-aminopurine source in the presence of 2'-deoxycoformycin, an inhibitor of adenine aminohydrolase activity. Although both knock-outs showed a diminished capacity to infect murine peritoneal macrophages, only the Δasl null mutant was profoundly incapacitated in its ability to infect mice. The enormous discrepancy in parasite loads observed in livers and spleens from mice infected with either Δadss or Δasl parasites can be explained by selective accumulation of adenylosuccinate in the Δasl knock-out and consequent starvation for guanylate nucleotides. Genetic complementation of a Δasl lesion in Escherichia coli implied that the L. donovani ASL could also recognize 5-aminoimidazole-(N-succinylocarboxamide) ribotide as a substrate, and purified recombinant ASL displayed an apparent Km of ∼24 μm for adenylosuccinate. Unlike many components of the purine salvage pathway of L. donovani, both ASL and ADSS are cytosolic enzymes. Overall, these data underscore the paramount importance of ASL to purine salvage by both life cycle stages of L. donovani and authenticate ASL as a potential drug target in Leishmania.
authors: Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, OR 97239, USA.
authors: Jan M
authors: JM
authors: Boitz
authors: Rona
authors: R
authors: Strasser
authors: Phillip A
authors: PA
authors: Yates
authors: Armando
authors: A
authors: Jardim
authors: Buddy
authors: B
authors: Ullman
mesh_terms: D000264:Adenylosuccinate Lyase / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D000265:Adenylosuccinate Synthase / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D000818:Animals; D001321:Autistic Disorder; D003001:Cloning, Molecular; D015195:Drug Design; D005260:Female; D005816:Genetic Complementation Test; D007700:Kinetics; D007893:Leishmania donovani / Q000235:genetics* / Q000472:pathogenicity* / Q000502:physiology; D007898:Leishmaniasis, Visceral / Q000188:drug therapy*; D008099:Liver / Q000378:metabolism / Q000469:parasitology; D008264:Macrophages / Q000166:cytology; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D016366:Open Reading Frames; D010641:Phenotype; D011686:Purine-Pyrimidine Metabolism, Inborn Errors / Q000235:genetics; D011687:Purines / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D011687:Purines; D012333:RNA, Messenger; D000264:Adenylosuccinate Lyase; D000265:Adenylosuccinate Synthase; C030985:purine
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