Biochemical characterization of the class B acid phosphatase (AphA) of Escherichia coli MG1655

Claudio Passariello, Costantino Forleo, Vanna Micheli, Serena Schippa, Rosalida Leone, Stefano Mangani, Maria Cristina Thaller, Gian Maria Rossolini

Research output: Contribution to journalArticlepeer-review


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.

Original languageEnglish
Pages (from-to)13-19
Number of pages7
JournalBiochimica et Biophysica Acta - Proteins and Proteomics
Issue number1
Publication statusPublished - Jan 2006


  • AphA
  • Biochemical characterization
  • class B acid phosphatase
  • Escherichia coli
  • Kinetic analysis

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Genetics


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