Enzymatic assay procedures that employ high-performance liquid chromatography: Competition between phosphoribosyltransferases for a common substrate

Donald L. Sloan, Linda Z. Ali, Diogenese Aybar-Batista, Cong Yan, Susan L. Hess

Research output: Contribution to journalArticle

5 Scopus citations

Abstract

A survey of the phosphoribosyltransferase (PRTase) activities in yeast has been accomplished using reversed-phase high-performance liquid chromatographic assay procedures. The following bases were observed to be utilized during phosphoribosyl pyrophosphate (PRibPP)-dependent nucleotide syntheses: adenine, xanthine, hypoxanthine, guanine, uracil, orotate, nicotinamide, nicotinate and quinolinate. Gradient elution procedures have also been perfected that allow the separation of the two following sets of PRTase assay components: (1) adenosine monophosphate, nicotinate mononucleotide, orotate, adenosine triphosphate, nicotinate, adenosine diphosphate, inosine monophosphate and hypoxanthine, and (2) nicotinate mononucleotide, nicotinamide mononucleotide, adenosine triphosphate, nicotinate, adenosine diphosphate and nicotinamide. Separation 1 has been employed to examine the PRibPP allocation among the hypoxanthine PRTase, orotate PRTase and nicotinate PRTase catalyzed reactions, whereas separation 2 has been employed to define the role that ATP plays in the nicotinamide PRTase-catalyzed reaction along with the allocation of nicotinamide between the reactions catalysed by nicotinamide PRTase and nicotinamide deamidase.

Original languageEnglish (US)
Pages (from-to)43-52
Number of pages10
JournalJournal of Chromatography A
Volume316
Issue numberC
DOIs
StatePublished - 1984
Externally publishedYes

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Organic Chemistry

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