Pathways of Pyrimidine Salvage in Pseudomonas and Former Pseudomonas: Detection of Recycling Enzymes Using High-Performance Liquid Chromatography

Abstract

Pyrimidine salvage pathways are vital for all bacteria in that they share in the synthesis of RNA with the biosynthetic pathway in pyrimidine prototrophs, while supplying all pyrimidine requirements in pyrimidine auxotrophs. Salvage enzymes that constitute the pyrimidine salvage pathways were studied in 13 members of Pseudomonas and former pseudomonads. Because it has been established that all Pseudomonas lack the enzyme uridine/cytidine kinase (Udk) and all contain uracil phosphoribosyl transferase (Upp), these two enzymes were not included in this experimental work. The enzymes assayed were: cytosine deaminase [Cod: cytosine + H2O --> uracil + NH3], cytidine deaminase [Cdd: cytidine + H2O --> uridine + NH3], uridine phosphorylase [Udp: uridine + Pi <--> uracil + ribose - 1 - P], nucleoside hydrolase [Nuh: purine/pyrimidine nucleoside + H2O --> purine/pyrimidine base + ribose], uridine hydrolase [Udh: uridine/cytidine + H2O --> uracil/cytosine + ribose]. The assay work generated five different Pyrimidine Salvage Groups (PSG) designated PSG1 - PSG5 based on the presence or absence of the five enzymes. These enzymes were assayed using reverse phase high-performance liquid chromatography techniques routinely carried out in our laboratory. Escherichia coli was included as a standard, which contains all seven of the above enzymes.