Abstract
Abstract The pyridine nucleotide content of wild-type Neurospora is 3 µmoles per g dry wt. It can be raised by 20% if cells are grown in the presence of nicotinic acid, and by 40% in the presence of tryptophan. This suggests that the utilization of tryptophan, the first step unique to pyridine nucleotide synthesis, is a rate-limiting step in this pathway. The pyridine nucleotide content of col-2, a morphological mutant containing a defective glucose 6-phosphate dehydrogenase, was only 40% of the wild-type strain. Growth of this mutant in the presence of nicotinic acid raised its pyridine nucleotide levels by approximately 20%. However, tryptophan had no effect on the levels in the col-2 strain. The lack of tryptophan stimulation in the col-2 strain suggests that there is a ratelimiting step elsewhere in the pyridine nucleotide pathway of this mutant. This step is postulated to be the one catalyzed by kynurenine hydroxylase, since this reaction requires NADPH and since the col-2 strain is deficient in NADPH. Studies on in vivo pyridine nucleotide turnover in a nicotinic acid-requiring strain demonstrated that the levels of NADH decreased faster than the levels of NADPH. By employing four different mutant strains and two double mutant strains grown under a variety of conditions, the total pyridine nucleotide content of Neurospora can be varied over approximately a 3-fold range, from 1.70 µmoles per g to 4.70 µmoles per g. Over this entire range, the ratio of the NAD nucleotides to NADP nucleotides remained at approximately 7:1. One of the factors which controlled this ratio was the level of the NADP type of nucleotide. Another ratio which was constant over the entire 3-fold range was the proportion of reduced pyridine nucleotides. The data indicate that the NADH/NAD + NADH ratio remained at 0.35 ± 0.03, and the NADPH/NADP + NADPH ratio remained at 0.65 ± 0.05. The implications of these constant oxidation-reduction ratios and the effects of the variations of the pyridine nucleotide content on the morphology of this organism are discussed.
Highlights
This paper reports the results of nicotinic acid and tryptophan supplementation on the pyridine nucleotide levels in wild-type Xeurospora, as well as the results obtained with four single and two double mutant strains blocked in different steps of pyridine nucleotide synthesis
The pyridine nucleotide content of the auxotrophic strain nit-S when grown with a supplement of nicotinic acid was the same as that of the wild type when grown on nicotinic acid (Table I)
The r&-S strain is alrnost completely devoid of kynurenine hydroxylase [4], and cannot synthesize its own pyridine nucleotides. It did not accumulate large amounts of pyridine nucleotides when the growth medium contains excess nicotinic acid, indicating that some rate-limiting step in the utilization of nicotinic acid prevented the overproduction of pyridine nucleotides
Summary
The pyridine nucleotide content of wild-type Neurospora is 3 kmoles per g dry wt It can be raised by 20% if cells are grown in the presence of nicotinic acid, and by 40% in the presence of tryptophan. The pyridine nucleotide content of ~01-2, a morphological mutant containing a defective glucose 6-phosphate dehydrogenase, was only 40 % of the wild-type strain Growth of this mutant in the presence of nicotinic acid raised its pyridine nucleotide levels by approximately 20%;. This paper reports the results of nicotinic acid and tryptophan supplementation on the pyridine nucleotide levels in wild-type Xeurospora, as well as the results obtained with four single and two double mutant strains blocked in different steps of pyridine nucleotide synthesis. To convert these values to average molar intracellular concentrations, one can assume that 1 g of dry mycelium occupies approximately 2.5 ml [6]
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