Cloning of a pyruvate phosphate dikinase from Trypanosoma cruzi

We have cloned and characterised a gene that encodes a putative pyruvate ph osphate dikinase (PPDK) from Trypanosoma cruzi, an enzyme that catalysts th e reversible conversion of phosphoenolpyruvate to pyruvate. PPDK is absent in mammalian cells, but has been Found in a wide variety of other organisms , including plants and bacteria. In T. cruzi, two genes (PPDK1 and PPDK2) a re present in a tandem array localised on a 1 Mbp chromosome. Northern and Western blot analyses indicates that PPDK is expressed as a 100-kDa protein in epimastigote. amastigote and trypomastigote forms. PPDK1 and PPDK2 enco de an identical protein of 100.8 kDa with a C-terminal extension ending wit h the sequence AKL, a signal for glycosomal import. Both T. cruzi and T. br ucei enzymes possess a 23-residue insertion, that is absent in other PPDKs. A three-dimensional alignment with the crystal structure of the: enzyme fr om Clostridium symbiosum predicts that this insertion is located on the sur face of the nucleotide-binding domain. Phylogenetic studies indicate that b acterial and protist PPDKs cluster as a separate group From those of plants . The evolutionary implications and possible role of this: enzyme in T cruz i is discussed. (C) 2001 Elsevier Science B.V. All rights reserved.