TcrPDEA1, a cAMP-specific phosphodiesterase with atypical pharmacological properties from Trypanosoma cruzi.
Cyclic nucleotide phosphodiesterases (PDEs) catalyze the degradation of cAMP and cGMP, and regulate a variety of cellular processes by controlling the levels of these second messengers. We have previously described the presence of both a calcium-stimulated adenylyl cyclase and two membrane-bound cAMP-specific PDEs (one of them strongly associated to the flagellum and the other one with a possible vesicular localization) in Trypanosoma cruzi. Here we report the identification and characterization of TcrPDEA1, a singular phosphodiesterase of T. cruzi which is resistant to the typical phosphodiesterase inhibitors, such as IBMX, papaverine and theofylline. TcrPDEA1 is a single copy gene that encodes a 620-amino acid protein, which is grouped with PDE1 family members, mainly with its kinetoplastid orthologs. TcrPDEA1 was able to complement a mutant yeast strain deficient in PDE genes, demonstrating that this enzyme is a functional phosphodiesterase. TcrPDEA1 is specific for cAMP with a high K(m) value (191.1+/-6.5 microM). Cyclic GMP neither activates the enzyme nor competes as a substrate. In addition, calcium-calmodulin did not affect the kinetic parameters and, as its counterpart in T. brucei, magnesium showed to be crucial for its activity and stability. Although TcrPDEA1 function remains unclear, its presence points out the high complexity of the cAMP signaling in trypanosomatids and the possible compartmentalization of the enzymes involved in the cAMP pathway.
Alonso
Guillermo D
GD
Consejo Nacional de Investigaciones Científicas y Técnicas and Departamento de Fisiología, Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina. galonso@dna.uba.ar
Schoijet
Alejandra C
AC
Torres
Héctor N
HN
Flawiá
Mirtha M
MM
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000494:pharmacology; D003067:Coenzymes / Q000494:pharmacology; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D020536:Enzyme Activators / Q000494:pharmacology; D004795:Enzyme Stability; D018628:Gene Dosage; D005816:Genetic Complementation Test; D006157:Guanosine Monophosphate / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D008969:Molecular Sequence Data; D010208:Papaverine / Q000494:pharmacology; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D013806:Theophylline / Q000494:pharmacology; D014349:Trypanosoma cruzi / Q000187:drug effects / Q000201:enzymology* / Q000254:growth & development
D002147:Calmodulin; D003067:Coenzymes; D016054:DNA, Protozoan; D020536:Enzyme Activators; D010726:Phosphodiesterase Inhibitors; D006157:Guanosine Monophosphate; D013806:Theophylline; D010208:Papaverine; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D008274:Magnesium; D002118:Calcium; D015056:1-Methyl-3-isobutylxanthine