Purification of escherichia coli B-specific p-aminobenzoate "pick-up" protein to homogeneity by affinity chromatography

Bela L. Toth-Martinez, Z. Dinya, Ferenc Hernádi

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Of the satellite fractions of Escherichia coli B dihydrofolate synthetases, a nonenzymic protein that is specifically able to bind p-aminobenzoate and sulphonamides has been purified 6000-fold by p-aminobenzoylcellulose affinity chromatography. The protein was named p-aminobenzoate "pick-up" protein according to its function, i.e., to bring p-aminobenzoate into reaction with l-glutamate and pteridine during dihydrofolate biosynthesis. About 4 mg of pure protein (0.532% recovery, calculated from the total p-aminobenzoate binding capacity of the unfractionated supernatant separated from the crude bacterium plasma) can be obtained from 500 g of harvested cells. The product is homogeneous in polyacrylamide gel electrophoresis both in the absence and presence of sodium dodecyl sulphate, and has a molecular weight of 15,000 daltons ±5% as measured by sodium dodecyl sulphate gel electrophoresis and Sephadex G-75 gel column chromatography. p-Aminobenzoate and sulphonamide ligand binding studies showed a single binding site per p-aminobenzoate pick-up protein molecule. KD values for p-aminobenzoate and some sulphonamides as well as for l-glutamate, l-γ-glutamyl oligopeptides, some pteridines and folate antagonists are also presented in order to illustrate the specificity of the receptor protein.

Original languageEnglish
Pages (from-to)205-211
Number of pages7
JournalJournal of Chromatography A
Volume115
Issue number1
DOIs
Publication statusPublished - Dec 10 1975

Fingerprint

para-Aminobenzoates
Affinity chromatography
Affinity Chromatography
Escherichia coli
Purification
Sulfonamides
Pteridines
Proteins
dihydrofolate synthetase
Electrophoresis
Sodium Dodecyl Sulfate
Glutamic Acid
Oligopeptides
Column chromatography
Biosynthesis
Folic Acid
Gel Chromatography
Polyacrylamide Gel Electrophoresis
Bacteria
Gels

ASJC Scopus subject areas

  • Analytical Chemistry
  • Clinical Biochemistry
  • Molecular Medicine

Cite this

Purification of escherichia coli B-specific p-aminobenzoate "pick-up" protein to homogeneity by affinity chromatography. / Toth-Martinez, Bela L.; Dinya, Z.; Hernádi, Ferenc.

In: Journal of Chromatography A, Vol. 115, No. 1, 10.12.1975, p. 205-211.

Research output: Contribution to journalArticle

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