title: Biochemical characterization of the class B acid phosphatase (AphA) of Escherichia coli MG1655.
abstract: The AphA enzyme of Escherichia coli, a molecular class B periplasmic phosphatase that belongs to the DDDD superfamily of phosphohydrolases, was purified and subjected to biochemical characterization. Kinetic analysis with several substrates revealed that the enzyme essentially behaves as a broad-spectrum nucleotidase highly active on 3'- and 5'-mononucleotides and monodeoxynucleotides, but not active on cyclic nucleotides, or nucleotides di- and triphosphate. Mononucleotides are degraded to nucleosides, and AphA apparently does not exhibit any nucleotide phosphomutase activity. However, it can transphosphorylate nucleosides in the presence of phosphate donors. Kinetic properties of AphA are consistent with structural data, and suggest a role for the hydrophobic pocket present in the active site crevice, made by residues Phe 56, Leu71, Trp77 and Tyr193, in conferring preferential substrate specificity by accommodating compounds with aromatic rings. AphA was inhibited by several chelating agents, including EDTA, EGTA, 1,10-phenanthroline and dipicolinic acid, with EDTA being apparently the most powerful inhibitor.
authors.affiliation: Dipartimento di Scienze di Sanità Pubblica, Università di Roma La Sapienza, 00185 Rome, Italy.
authors.forename: Claudio
authors.initials: C
authors.lastname: Passariello
authors.forename: Costantino
authors.initials: C
authors.lastname: Forleo
authors.forename: Vanna
authors.initials: V
authors.lastname: Micheli
authors.forename: Serena
authors.initials: S
authors.lastname: Schippa
authors.forename: Rosalida
authors.initials: R
authors.lastname: Leone
authors.forename: Stefano
authors.initials: S
authors.lastname: Mangani
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Thaller
authors.forename: Gian Maria
authors.initials: GM
authors.lastname: Rossolini
mesh_terms: D000135:Acid Phosphatase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D020134:Catalytic Domain; D002614:Chelating Agents / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008958:Models, Molecular; D009711:Nucleotides / Q000378:metabolism; D020836:Protein Structure, Quaternary; D013379:Substrate Specificity; D013816:Thermodynamics
chemical_list: D002614:Chelating Agents; D004791:Enzyme Inhibitors; D009711:Nucleotides; D000135:Acid Phosphatase