Publications

• Human pyridoxal phosphatase. Molecular cloning, functional expression, and tissue distribution.

  Jang Y.M., Kim D.W., Kang T.-C., Won M.H., Baek N.-I., Moon B.J., Choi S.Y., Kwon O.-S.

  Pyridoxal phosphatase catalyzes the dephosphorylation of pyridoxal 5'-phosphate (PLP) and pyridoxine 5'-phosphate. A human brain cDNA clone was identified to the PLP phosphatase on the basis of peptide sequences obtained previously. The cDNA predicts a 296-amino acid protein with a calculated Mr o ... >> More

  Pyridoxal phosphatase catalyzes the dephosphorylation of pyridoxal 5'-phosphate (PLP) and pyridoxine 5'-phosphate. A human brain cDNA clone was identified to the PLP phosphatase on the basis of peptide sequences obtained previously. The cDNA predicts a 296-amino acid protein with a calculated Mr of 31698. The open reading frame is encoded by two exons located on human chromosome 22q12.3, and the exon-intron junction contains the GT/AG consensus splice site. In addition, a full-length mouse PLP phosphatase cDNA of 1978 bp was also isolated. Mouse enzyme encodes a protein of 292 amino acids with Mr of 31512, and it is localized on chromosome 15.E1. Human and mouse PLP phosphatase share 93% identity in protein sequence. A BLAST search revealed the existence of putative proteins in organism ranging from bacteria to mammals. Catalytically active human PLP phosphatase was expressed in Escherichia coli, and characteristics of the recombinant enzyme were similar to those of erythrocyte enzyme. The recombinant enzyme displayed Km and kcat values for pyridoxal of 2.5 microM and 1.52 s(-1), respectively. Human PLP phosphatase mRNA is differentially expressed in a tissue-specific manner. A single mRNA transcript of 2.1 kb was detected in all human tissues examined and was highly abundant in the brain. Obtaining the molecular properties for the human PLP phosphatase may provide new direction for investigating metabolic pathway involving vitamin B6. << Less

  J. Biol. Chem. 278:50040-50046(2003) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Kinetic mechanism and divalent metal activation of human erythrocyte pyridoxal phosphatase.

  Fonda M.L., Zhang Y.N.

  Human erythrocyte pyridoxal phosphatase has an essential requirement for divalent cations. Its activation by Mg2+, Co2+, Ni2+, or Mn2+ followed Michaelis-Menten kinetics. Other divalent cations inhibited the enzyme. The kinetic properties of the enzyme were investigated with pyridoxal phosphate an ... >> More

  Human erythrocyte pyridoxal phosphatase has an essential requirement for divalent cations. Its activation by Mg2+, Co2+, Ni2+, or Mn2+ followed Michaelis-Menten kinetics. Other divalent cations inhibited the enzyme. The kinetic properties of the enzyme were investigated with pyridoxal phosphate and Mg2+ alone and in the presence of the product, Pi, or dead-end inhibitors at pH 7.4 and 37 degrees C. The enzyme bound both the substrate and Mg2+ before products were released. Pi gave competitive inhibition vs substrate and noncompetitive inhibition vs Mg2+. Molybdate also was a competitive inhibitor vs substrate and noncompetitive inhibitor vs Mg2+. Ca2+ gave competitive inhibition vs Mg2+ and noncompetitive inhibition vs substrate. The effects of Mg2+ and substrate on the inactivation of pyridoxal phosphatase by a variety of group-specific reagents were studied. The inactivation of the enzyme by iodoacetate was potentiated by MgCl2. The Kd of the enzyme-Mg complex determined in the inactivation analysis was similar to the Km of the free enzyme for Mg2+, indicating that Mg2+ binds to the free enzyme. Low concentrations of a substrate, pyridoxine phosphate, or Pi protected pyridoxal phosphatase from inactivation by N-ethylmaleimide in the absence or presence of Mg2+. Thus, the substrate binds to the free enzyme and the enzyme-Mg complex. The steady-state kinetics and the kinetics of inactivation are consistent with random binding of pyridoxal phosphate and Mg2+ and with the formation of a dead-end complex of Pi with the enzyme-Mg complex. << Less

  Arch Biochem Biophys 320:345-352(1995) [PubMed] [EuropePMC]

• Purification and characterization of vitamin B6-phosphate phosphatase from human erythrocytes.

  Fonda M.L.

  Human erythrocytes rapidly convert vitamin B6 to pyridoxal-P and contain soluble phosphatase activity which dephosphorylates pyridoxal-P at a pH optimum of 6-6.5. This phosphatase was purified 51,000-fold with a yield of 39% by ammonium sulfate precipitation and chromatography on DEAE-Sepharose, S ... >> More

  Human erythrocytes rapidly convert vitamin B6 to pyridoxal-P and contain soluble phosphatase activity which dephosphorylates pyridoxal-P at a pH optimum of 6-6.5. This phosphatase was purified 51,000-fold with a yield of 39% by ammonium sulfate precipitation and chromatography on DEAE-Sepharose, Sephacryl S-200, hydroxylapatite, and reactive yellow 86-agarose. Sephacryl S-200 chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the enzyme was a dimer with a molecular mass of approximately 64 kDa. The phosphatase required Mg2+ for activity. It specifically catalyzed the removal of phosphate from pyridoxal-P, pyridoxine-P, pyridoxamine-P, 4-pyridoxic acid-P, and 4-deoxypyridoxine-P at pH 7.4. Nucleotide phosphates, phosphoamino acids, and other phosphorylated compounds were not hydrolyzed significantly nor were they effective inhibitors of the enzyme. The phosphatase showed Michaelis-Menten kinetics with its substrates. It had a Km of 1.5 microM and a Vmax of 3.2 mumol/min/mg with pyridoxal-P. The Vmax/Km was greatest with pyridoxal-P greater than 4-pyridoxic acid-P greater than pyridoxine-P greater than pyridoxamine-P. The phosphatase was competitively inhibited by the product, inorganic phosphate, with a Ki of 0.8 mM, and weakly inhibited by pyridoxal. It was also inhibited by Zn2+, fluoride, molybdate, and EDTA, but was not inhibited by levamisole, L-phenylalanine, or L(+)-tartrate. These properties of the purified enzyme suggest that it is a unique acid phosphatase that specifically dephosphorylates vitamin B6-phosphates. << Less

  J Biol Chem 267:15978-15983(1992) [PubMed] [EuropePMC]

• Pancreatitis with hyperlipaemic serum occurring with hypogonadism and congential blindness.

  Koelmeyer T.D., Sorrell V.F.

  N Z Med J 79:693-695(1974) [PubMed] [EuropePMC]

• Probing the substrate specificities of human PHOSPHO1 and PHOSPHO2.

  Roberts S.J., Stewart A.J., Schmid R., Blindauer C.A., Bond S.R., Sadler P.J., Farquharson C.

  PHOSPHO1, a phosphoethanolamine/phosphocholine phosphatase, is upregulated in mineralising cells and is thought to be involved in the generation of inorganic phosphate for bone mineralisation. PHOSPHO2 is a putative phosphatase sharing 42% sequence identity with PHOSPHO1. Both proteins contain thr ... >> More

  PHOSPHO1, a phosphoethanolamine/phosphocholine phosphatase, is upregulated in mineralising cells and is thought to be involved in the generation of inorganic phosphate for bone mineralisation. PHOSPHO2 is a putative phosphatase sharing 42% sequence identity with PHOSPHO1. Both proteins contain three catalytic motifs, conserved within the haloacid dehalogenase superfamily. Mutation of Asp32 and Asp203, key residues within two motifs, abolish PHOSPHO1 activity and confirm it as a member of this superfamily. We also show that Asp43 and Asp123, residues that line the substrate-binding site in our PHOSPHO1 model, are important for substrate hydrolysis. Further comparative modelling reveals that the active sites of PHOSPHO1 and PHOSPHO2 are very similar, but surprisingly, recombinant PHOSPHO2 hydrolyses phosphoethanolamine and phosphocholine relatively poorly. Instead, PHOSPHO2 shows high specific activity toward pyridoxal-5-phosphate (V(max) of 633 nmol min(-1) mg(-1) and K(m) of 45.5 microM). Models of PHOSPHO2 and PHOSPHO1 suggest subtle differences in the charge distributions around the putative substrate entry site and in the location of potential H-bond donors. << Less

  Biochim. Biophys. Acta 1752:73-82(2005) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.