Genetic Characterization of Pyridine Nucleotide Uptake Mutants of Salmonella typhimurium

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Abstract
Two classes of pyridine nucleotide uptake mutants isolated previously in a strain of S. typhimurium defective in both de novo NAD biosynthesis (nad) and pyridine nucleotide recycling (pncA) were analyzed in terms of their genetic relationship to each other and their roles in the transport of nicotinamide mononucleotide (NMN) as a precursor to NAD. The 1st class of uptake mutants, pnuA (99 units), failed to grow on NMN as a precursor for NAD. The 2nd class, pnuB, grew on lower than normal levels of NMN and suppressed pnuA mutations. A 3rd class of uptake mutant, pnuC, isolated in a nadB pncA pnuB background, also failed to grow on NMN. Transport studies and enzyme analyses confirmed these strains as defective in NMN uptake. A 4th locus, designated pnuD, was found to diminish NMN utilization in a nad pncA+ background. Tn10 insertions near pnuA, pnuC and pnuD were isolated and utilized in mapping studies. pnuA was found to map between thr and serB near trpR. The pnuC locus was cotransducible with nadA at 17 units while pnuD mapped at .apprx. 60 units. The biochemical and genetic data suggest that the pnuA and pnuC gene products cooperate in the utilization of NMN under normal conditions. A pnuB mutant, however, does not require the pnuA gene product for NMN uptake but does rely on the pnuC product. Fusion studies indicate that pnuC is regulated by internal NAD concentrations.

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