title: Overexpression of a Zn2+-sensitive soluble exopolyphosphatase from Trypanosoma cruzi depletes polyphosphate and affects osmoregulation.
abstract: We report the cloning, expression, purification, and characterization of the Trypanosoma cruzi exopolyphosphatase (TcPPX). The product of this gene (TcPPX), has 383 amino acids and a molecular mass of 43.1 kDa. TcPPX differs from most exopolyphosphatases in its preference for short-chain polyphosphate (poly P). Heterologous expression of TcPPX in Escherichia coli produced a functional enzyme that had a neutral optimum pH and was dramatically inhibited by low concentrations of Zn2+, high concentrations of basic amino acids (lysine and arginine), and heparin. TcPPX is a processive enzyme and does not hydrolyze ATP, pyrophosphate, or p-nitrophenyl phosphate, although it hydrolyzes guanosine 5'-tetraphosphate very efficiently. Overexpression of TcPPX resulted in a dramatic decrease in total short-chain poly P and partial decrease in long-chain poly P. This was accompanied by a delayed regulatory volume decrease after hyposmotic stress. These results support the role of poly P in T. cruzi osmoregulation.
authors.affiliation: Center for Tropical and Emerging Global Diseases and Department of Cellular Biology, University of Georgia, Paul D. Coverdell Biomedical and Health Sciences Center, Athens, Georgia 30602, USA.
authors.forename: Jianmin
authors.initials: J
authors.lastname: Fang
authors.forename: Felix A
authors.initials: FA
authors.lastname: Ruiz
authors.forename: Melissa
authors.initials: M
authors.lastname: Docampo
authors.forename: Shuhong
authors.initials: S
authors.lastname: Luo
authors.forename: Juliany C F
authors.initials: JC
authors.lastname: Rodrigues
authors.forename: Lucimar S
authors.initials: LS
authors.lastname: Motta
authors.forename: Peter
authors.initials: P
authors.lastname: Rohloff
authors.forename: Roberto
authors.initials: R
authors.lastname: Docampo
mesh_terms: D017766:Acid Anhydride Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D008969:Molecular Sequence Data; D011122:Polyphosphates / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017385:Sequence Homology; D014349:Trypanosoma cruzi / Q000201:enzymology*; D014882:Water-Electrolyte Balance; D015032:Zinc / Q000378:metabolism*