Open AccessNucleotide phosphotransferase of Escherichia coli: purification by affinity chromatography.
Author(s) -
Elinor F. Brunngraber,
Erwin Chargaff
Publication year - 1977
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.74.8.3226
Subject(s) - escherichia coli , nucleotide , biochemistry , affinity chromatography , agarose , phosphotransferase , chemistry , enzyme , cytidine , dna , pep group translocation , phosphate , chromatography , microbiology and biotechnology , biology , gene
Improved extraction and purification procedures permit the isolation from Escherichia coli W cells of much larger quantities and of more highly purified preparations of nucleotide phosphotransferase. Of various affinity resins tested for efficiency of purification, columns of agarose/5'-AMP (AGAMP), type 3, proved the best. In this way a 300- to 450-fold purification of the enzyme was achieved in a few steps. The enzyme, which, as reported before, transfers organically bound phosphate to the 2' or 3' hydroxyls of nucleosides and nucleotides, was tested in its behavior toward a series of ribonucleosidonucleotides, namely, CpC, ApA, CpA, and ApC. All were phosphate acceptors, but a detailed comparative study of adenosine and cytidine, 5'-AMP and 5'-CMP, and ApA and ApC revealed peculiar specificities in the relative distribution of the phosphorylated products.