Trypanosoma cruzi arginine kinase characterization and cloning. A novel energetic pathway in protozoan parasites.
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.
Pereira
C A
CA
Instituto de Investigaciones en Ingeniería Genética y Biología Molecular, Consejo Nacional de Investigaciones Científicas y Técnicas, and Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina.
Alonso
G D
GD
Paveto
M C
MC
Iribarren
A
A
Cabanas
M L
ML
Torres
H N
HN
Flawiá
M M
MM
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D001122:Arginine Kinase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D002413:Cations, Divalent / Q000494:pharmacology; D002846:Chromatography, Affinity; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D015698:Genomic Library; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014349:Trypanosoma cruzi / Q000201:enzymology* / Q000235:genetics*
D000596:Amino Acids; D002413:Cations, Divalent; D011994:Recombinant Proteins; D001122:Arginine Kinase