Histidine regulation in Salmonella typhimurium. XVI. A sensitive radiochemical assay for histidinol dehydrogenase

Zygmunt Ciesla, Francesco Salvatore, James R. Broach, Stanley W. Artz, Bruce N. Ames

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

Abstract

A sensitive radiochemical assay for measurement of histidinol dehydrogenase is presented. The method is based upon separation of the product of the reaction. [14C]histidine, from the substrate, [14C]histidinol, on small Dowex 50 columns. The assay can be performed on cell extracts or on toluenized cells and is approximately 100 times more sensitive than previously reported assays for this enzyme. [14C]histidinol is obtained in high yields through conversion of uniformly labeled 14C-glucose by a strain of Salmonella typhimurium derepressed for the histidine operon and blocked at the histidinol dehydrogenase step. Accumulated [14C]histidinol is purified from the culture supernatant by ion-exchange chromatography. This sensitive assay has facilitated measurement of reduced levels of histidine operon expression in promoter mutants, and has been adapted for study of histidine operon regulation in a cell free protein synthesizing system.

Original languageEnglish (US)
Pages (from-to)44-55
Number of pages12
JournalAnalytical Biochemistry
Volume63
Issue number1
DOIs
StatePublished - Jan 1975

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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