Deoxycytidine triphosphate deaminase: identification and function in Salmonella typhimurium

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• The biosynthesis of 2'-deoxyuridine monophosphate (dUMP) is crucial for DNA synthesis.
• Understanding dUMP metabolic pathways in bacteria like Salmonella typhimurium provides insights into nucleotide metabolism.

Purpose of the Study:

• To investigate the biosynthesis pathways of dUMP in a Salmonella typhimurium mutant.
• To elucidate the roles of uridine and cytosine precursors in dUMP synthesis.
• To analyze the impact of thymine starvation on dUMP pools and metabolism.

Main Methods:

• Utilized a cytidine- and uracil-requiring Salmonella typhimurium mutant (DP-55).
• Traced dUMP and thymidine monophosphate (dTMP) synthesis using radiolabeled uracil and unlabeled cytidine.
• Identified and partially purified Mg(2+)-dependent 2'-deoxycytidine triphosphate (dCTP) deaminase.
• Introduced a thymine requirement to study effects on dUMP pools and catabolism.

Main Results:

• DP-55 mutant primarily synthesized dUMP (70%) from cytosine compounds, with only 30% from uridine.
• dCTP deaminase and dUTP pyrophosphatase were identified as key enzymes in the cytosine-derived dUMP synthesis pathway.
• Introducing a thymine requirement led to a 100-fold increase in the dUMP pool and augmented dUMP catabolism to uracil.
• Partial thymine starvation increased dUMP and dCTP pools, enhancing the contribution of the dCTP pathway to dUMP synthesis.

Conclusions:

• The primary route for dUMP biosynthesis in Salmonella typhimurium involves a cytosine-derived pathway, mediated by dCTP deaminase and dUTP pyrophosphatase.
• The dUMP pool size is tightly regulated, with significant catabolism to uracil occurring under conditions of high dUMP levels or thymine starvation.
• These findings highlight the complex interplay between nucleotide salvage and de novo synthesis pathways in bacterial DNA precursor metabolism.