Trypanosoma cruzi arginine kinase characterization and cloning. A novel energetic pathway in protozoan parasites

• Published in: The Journal of biological chemistry 2000-01, Vol.275 (2), p.1495-1501
• Main Authors: Pereira, C A, Alonso, G D, Paveto, M C, Iribarren, A, Cabanas, M L, Torres, H N, Flawiá, M M
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino Acids - pharmacology; Animals; arginine; Arginine Kinase - genetics; Arginine Kinase - isolation & purification; Arginine Kinase - metabolism; Base Sequence; canavanine; Cations, Divalent - pharmacology; Chromatography, Affinity; Chromosome Mapping; Cloning, Molecular; Genomic Library; histidine; Humans; Kinetics; Molecular Sequence Data; Phylogeny; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Sequence Alignment; Sequence Homology, Amino Acid; Trypanosoma cruzi; Trypanosoma cruzi - enzymology; Trypanosoma cruzi - genetics
• ISSN: 0021-9258
• DOI: 10.1074/jbc.275.2.1495

This work contains the first description of a guanidino kinase in a flagellar unicellular parasite. The enzyme phosphorylates L-arginine and was characterized in preparations from Trypanosoma cruzi, the ethiological agent of Chagas' disease. The activity requires ATP and a divalent cation. Under standard assay conditions (1 mM L-arginine), the presence of 5-fold higher concentrations of canavanine or histidine produced a greater than 50% enzyme inhibition. The base sequence of this enzyme revealed an open reading frame of 357 amino acids and a molecular weight of 40,201. The amino acid sequence shows all of the characteristic consensus blocks of the ATP:guanidino phosphotransferase family and a putative "actinin-type" actin-binding domain. The highest amino acid identities of the T. cruzi sequence, about 70%, were with arginine kinases from Arthropoda. Southern and chromosome blots revealed that the kinase is encoded by a single-copy gene. Moreover, Northern blot analysis showed an mRNA subpopulation of about 2.0 kilobases, and Western blotting of T. cruzi-soluble polypeptides revealed a 40-kDa band. The finding in the parasite of a phosphagen and its biosynthetic pathway, which are totally different from those in mammalian host tissues, points out this arginine kinase as a possible chemotherapy target for Chagas' disease.