Effect of Histidine on Purine Nucleotide Synthesis and Utilization in Neurospora crassa

Transcription

1 JOURNAL OF BACTERIOLOGY, Oct. 1975, p Copyright 1975 American Society for Microbiology Vol. 124, No. 1 Printed in U.S.A. Effect of Histidine on Purine Nucleotide Synthesis and Utilization in Neurospora crassa LAKSHMI PENDYALA AND ANGELA M. WELLMAN* Department of Plant Sciences, The University of Western Ontario, London, Ontario, Canada Received for publication 7 July 1975 Histidine affects de novo purine nucleotide synthesis and purine nucleotide pool utilization in Neurospora crassa. The former effect was assessed qualitatively by the presence or absence of purple pigment production in ad3b and ad3a mutants. Tryptophan also affected the de novo purine nucleotide synthesis. The effect of histidine on purine nucleotide pool utilization resulted in stimulated germination of ad8 and ad4 mutant conidia in adenine-deficient medium. Increased germination was correlated with increased net levels of nucleic acids in these strains. Possible mechanisms for the dual action of histidine are discussed. From the studies on Escherichia coli purinerequiring mutants, a cyclical interaction between adenine and histidine pathways has been described by Magasanik and Karibian (13) and Sheldovsky and Magasanik (16). The main features of this interaction are that (i) adenosine triphosphate (ATP) is a precursor for histidine biosynthesis and (ii) a side product of histidine biosynthesis, amino imidazole carboxamide ribonucleotide (AICAR), is an intermediate in adenine nucleotide biosynthesis. The interaction between the two pathways produces different consequences for adenine mutants as far as the nucleotide pool utilization is concerned. In those adenine mutants that are blocked after AICAR formation in adenine nucleotide biosynthesis, part of the adenine nucleotide pool spent on histidine biosynthesis is wasted as nonusable AICAR. In the presence of histidine, however, this expenditure of the adenine nucleotide pool for histidine synthesis is spared, since histidine exerts a feedback regulation on the enzyme that uses ATP as a substrate. In Neurospora, Ahmed (Ph.D. thesis, Yale Univ., New Haven, Conn., 1964) has shown that histidine regulates the activity of Nl-(5'- phosphoribosyl)atp:pyrophosphate phosphoribosyltransferase (EC ), the first enzyme unique to the histidine pathway, by feedback inhibition. Therefore, histidine can be expected to exert a sparing effect on the nucleotide pool utilization in the adenine mutants of Neurospora blocked after AICAR formation in purine nucleotide biosynthesis. There is some indirect evidence available in Neurospora to show that this may be the case (12; T. Ishikawa, Ph.D. thesis, Yale Univ., ). We present additional evidence that histidine indeed has a sparing effect on Neurospora adenine mutants located in the terminal steps of the pathway. Figure 1 shows the adenine pathway and its likely relationship with the histidine pathway. The locations of mutational blocks are also shown for all the available Neurospora adenine mutants as demonstrated by Bernstein (4), Buchanan (5), and Jha (9). When the conidia of different adenine mutants of Neurospora are incubated on adeninedeficient medium, some mutants germinate readily whereas the others do not. The mutant conidia that do not germinate readily fall into the later steps of the pathway, specifically after AICAR formation. It has been shown with four selected adenine mutants that inosine forms the bulk of the conidial storage reserves, and adenine nucleotides constitute most of the remainder of the pool (15). Two of the mutants used in the above study were ad8 and ad4, whose mutational blocks fall in the very last two steps of adenine nucleotide biosynthesis; thus they are both AICAR and inosine monophosphate nonutilizers. The poor germination of these mutant conidia, despite their endogenous nucleotide reserves, indicated a possible drainage of some of the reserves to histidine synthesis. Therefore, the reserve purine pool utilization was studied in these mutants in the presence and absence of histidine. Involvement of histidine in a cross-pathway regulation of enzymes concerned with histidine, tryptophan, and arginine biosynthesis was recently demonstrated in Neurospora (6). We report here that in addition to influencing the Downloaded from on October 19, 218 by guest

2 VOL. 124, 1975 PURINE METABOLISM IN N. CRASSA 79 5 P- Ribosyl Pyrophosphate (PRPP) Glutomine ~~~~~~Anthranilic Acid 5P-R bos ATP 5P-Ribosylamine (PRA) N(5'-Phosphoribosyl)- I AnthranilicAcid 5P-Rlbosyl AMP Glycinamide-RP (GAR) *~~~~~~~~~tad9 otn-formylglycinamide-rp (FGAR) I t~~~~~~~~ ad6 Imidozol Glycerol Phosphote Formyl Glycinamidine-RP(FGAM) Indole-3-Glycerol.-I ad 2 Phosphate Dl Imidazole Acetol Phosphate 5-Amino Imidozole-PP(AIR) Tryptophan Histidinol Phosphate ad 3B 5-Amino lmidozole Carboxylate-RP(CAIR) i Hstidinol IHistidinol ad 5-Amino Imidazole 3A 4-N Succino Carbox- mide-rp(saicar) L- ~~~~~~~~~~~ad4 Histidinel 5-Amino Imidazole Corboxcamide-RP(AICAR) P ad l or ad5 Formyl Amino Imidazole-RP(FAICAR) # ad 5 or od 1 Inosine-P 'IMP) Adenylosuccinic Acid(AMP-S) Xanthylic Acid (XMP) *ad4 i Inosine - Adenylic Acid (AMP) Guanylic Acid (GMP) ADP Exogenous Adenine -Ribonucleic Acid I -Other Metabolic Reactions FIG. 1. Adenine biosynthetic pathway, location of the mutational blocks of adenine mutants of N. crassa, and the interrelationships of adenine, histidine, and tryptophan pathways. pool utilization in some adenine mutants, histidine also has an effect on the de novo adenine nucleotide synthesis. The possible mechanisms by which histidine and tryptophan might influence the de novo adenine nucleotide synthesis will be discussed. MATERIALS AND METHODS The origin and maintenance of stocks, germination tests, preparation of labeled conidia, and pool utilization studies were as described previously (15). Histidine (2.5 mm) was used for germination and pool utilization studies. L-Histidine, L-tryptophan, L-alanine, L-arginine, L-glutamic acid, adenine sulfate, and uracil were purchased from Nutritional Biochemicals Corp., Cleveland, Ohio. Glycine and L-proline were purchased from Sigma Chemical Co., St. Louis, Mo C]adenine was purchased from ICN Chemical and Radioisotope Division, Irvine, Calif. Purple pigment production in ad3b ad3a mutants. In Fries liquid minimal medium containing up to 5Mtg of adenine per ml, ad3b and ad3a mutants grow slowly and produce a purple pigment. In ad3b and ad3a mutants the effect of various amino acids and uracil on pigment production was tested by supplementing them at 5 Mg/ml, in the presence of 5 Ag of adenine per ml. RESULTS Germination responses of mutant conidia on histidine-supplemented medium. The conidia for germination tests were obtained from mutant cultures grown on 2 mm adenine-supplemented medium. The conidia were spread on either histidine-supplemented medium or minimal medium (controls) and incubated for 6 h at 3 C. The results of these germination tests are shown in Table 1. The histidine supplementation stimulated the average percentage of germination in ad8 and ad4 strains, but the level of germination did not approach that of other mutants, such as ad9 and ad3b. Effect of histidine on the purine pool utilization. IC-labeled conidia of four adenine mutants and wild type were obtained from growth on 2 mm ["C ]adenine-supplemented agar medium (specific activity,.1 Ci/mol). These conidia had all their adenine-derived fractions (nucleic acids and acid-soluble pools) labeled. Spore suspensions of these labeled conidia were prepared in minimal medium or unlabeled histidine-supplemented medium to give 6 x 16 spores/ml and shaken at 3 C for 1 TABLE 1. Effect of histidine on the germination of conidia in some adenine mutants of Neurospora Avg % germinationa Locus Allele no. Minimal + Minimal histidine ad9 Y154M37A 81.5 (±1.) 79.6 (±2.) ad3b (±7.) 78.9 (±3.) ad8 Y112M (+4.) 36.2 (4-1.) ad4 Y152M (+7.5) 55. (±5.3) a Average of two different experiments. Spores in 16 to 24 fields (x4 objective) were counted for each strain in each experiment. Standard deviations are given in parentheses. Downloaded from on October 19, 218 by guest

3 8 PENDYALA AND WELLMAN to 11 h. From the samples taken at hourly intervals, the total radioactivity in the conidia and the radioactivity remaining after cold 5% trichloroacetic acid extraction (the radioactivity incorporated into nucleic acid) was determined by counting in 15 ml of scintillation fluid containing 2,5-diphenyloxazole and 1,4-bis- [2]-(5-phenyloxazolyl)benzene in toluene in a Beckman scintillation spectrometer. The acidsoluble pool is the difference between the total and acid-insoluble fractions. Figure 2 shows the J. BACTOL. percentage of the initial total radioactivity in the conidia at'each sampling time, when labeled conidia of wild-type and mutant strains were incubated in minimal or histidine media. The total radioactivity in the conidia of different strains at zero time ranged from 6,8 to 1,3 counts/min per sample. As seen in Fig. 2 and reported earlier (15), the conidia of all the strains, except ad9, leak significant amounts of a labeled compound when incubated in minimal medium. Incubation in histidine-supple- A z ad 9 p * * I s lime (HOURS) ad 3B Downloaded from on October 19, 218 by guest FIG. 2. Effect of histidine on the leakage of label from conidia. "4C-labeled conidia were suspended in unlabeled minimal or histidine-supplemented medium, and the radioactivity associated with each sample at hourly intervals was determined. Percentage of the initial total radioactivity present in conidia at each sampling period is presented against time. Symbols: () Conidia incubated in minimal medium; (A) conidia incubated in histidine-supplemented medium.

4 VOL. 124, 1975 mented medium had not altered the leakage pattern in wild-type, ad9, and ad3b strains. However, incubation of ad8 and ad4 conidia in histidine-supplemented medium resulted in a partial sparing of the leakage of the labeled compound. Some leakage still occurred in the first 6 h, but after 6 h in the presence of PURINE METABOLISM IN N. CRASSA 81 histidine the loss of label stopped in both the strains. Figure 3 shows the utilization of endogenous purine pools for nucleic acid synthesis in wild type and four adenine mutants in minimal medium and in the presence of histidine. In all those strains where reserve purine pools were x E <I 3 U C t- O 1 x13 Q ou 6J OL >_ 21 Qu I- Q 4 13 U3 CK 4- U WILD TYPE \\\ "\NA _ -, 4-'. A_ " M 1 TIME(HOURS) ad9 INS 1-11'6,4 & A _ - _I- -I TIME( HOURS) - ad8 -& - b --A ---o x13 E I- Q. U z ~-3 u - xl r E u 4 4 I- >73 I-- U < 2 4 K ad3 B \ _-8 -~~~~ -8 oss ^ ~ a TIME(HOURS) A \ " ad4 1 _A - o 4--a -- -d-_ 4- Downloaded from on October 19, 218 by guest U 2 4 b TIME(HOURS) TIME (HOURS) FIG. 3. Effect of histidine on the utilization of purine pools for nucleic acid synthesis. "4C-labeled conidia were suspended in unlabeled minimal or histidine-supplemented medium, and the radioactivity associated with the acid-insoluble and acid-soluble fractions was determined at hourly intervals. Symbols: () Conidia incubated in minimal medium; (A) conidia incubated in histidine-supplemented medium. Solid line, Acid-insoluble fraction; dotted line, acid-soluble fraction.

5 82 PENDYALA AND WELLMAN utilized for nucleic acid synthesis, a net increase in nucleic acid content was observed for up to 6 h, followed by a characteristic decline. This characteristic increase in nucleic acid and turnover at 6 h was also observed in all strains when conidia were incubated in adenine medium (15). The changes in the acid-soluble pools correspond to the amounts contributed for nucleic acid synthesis and/or leakage in various strains. For wild type, ad9, and ad3b the amount of increase and turnover in "4C-labeled nucleic acid was the same in minimal or histidine-supplemented media. ad8 and ad4 strains showed a net decrease of nucleic acid starting after 3 to 4 h of incubation in minimal medium. When incubated in histidine-supplemented medium, 75% of this nucleic acid breakdown was spared in ad8 and no net loss in labeled nucleic acid was found for ad4. Effect of histidine on the purple pigment production of ad3b and ad3a mutants. The structure of the characteristic purple pigment produced in ad3b and ad3a mutants of Neurospora is not known. Mitchell and Houlahan (14) suggested that "the pigment arises as a metabolic product of a labile adenine precursor, that accumulates as a result of the genetic block in adenine synthesis." In Saccharomyces the adenine mutants adl and ad2, which correspond to ad3b and ad3a mutants of Neurospora, also accumulate a red pigment. In this organism, the pigment was identified as a mixture of poly(ribosyl aminoimidazole) molecules varying in molecular weight and containing a number of amino acids (17). In Saccharomyces the pigment production of adl and ad2 mutants was reduced by exogenous adenine. Armitt and Woods (1) isolated mutants defective in the regulation of de novo purine synthesis that continued to accumulate pigment in the presence of excess adenine. Applying the same argument to the current work we can reason that, since intense pigment formation was observed only in low adenine concentrations, its production must represent a continued de novo purine synthetic activity. In high adenine concentrations the pigment does not form. It has been shown in many systems that de novo purine synthesis is inhibited when cells are incubated with high concentrations of purine bases (7). The first enzyme unique to purine nucleotide synthesis, amido phosphoribosyltransferase (EC ), was believed to be the target enzyme for feedback inhibition (2). Alternatively, Bagnara et al. (3), working with Ehrlich ascites tumor cells, proposed that lowering of intracellular phosphoribosyl pyrophosphate (PRPP) concentration is the cause of inhibition of de novo purine synthesis. Whatever may be the mechanism, in Neurospora de novo purine synthesis appears to be inhibited at high adenine concentrations but not at concentrations up to 5 gg/ml, as demonstrated by purple pigment formation in the ad3b and ad3a mutants. In the present study two different alleles of ad3b mutants and two different alleles of ad3a mutants were used to assess the de novo purine synthetic ability of N. crassa in the presence of externally supplied histidine, other amino acids, and uracil (Table 2). In the first 3 days of growth, histidine and tryptophan inhibited purple pigment formation in all ad3b and ad3a strains tested. After this period the mutants began producing the pigment. From the dry weights of these mutants, taken at two different periods (Table 3), it appears that inhibition of pigment formation is not a secondary effect due to growth inhibition for cultures grown in histidine-containing medium. In fact, the dry weights of mutants taken at 12 h showed a slight stimulation of growth in this medium. Mutants grown in tryptophan-containing media, however, showed a reduction in growth at both periods. Whether lag in pigment formation observed in tryptophan medium can be correlated with the observed reduction in growth in this medium is not clear. Interestingly, all tryptophan-grown cultures including wild type TABLE 2. Purple pigment production in ad3b and ad3a mutants in the presence of different amino acids and uracil Minimal medium supplemented with:" J. BACTERIOL. Purple pigment production in: ad3b ad3b ad3a ad3a Adenine Adenine + glycine Adenine + proline Adenine + alanine Adenine + arginine Adenine + glutamic acid Adenine + histidine Adenine + tryptophan Adenine + uracil asupplementation with all the amino acids and uracil was 5 ug/ml; supplementation with adenine was 5 gg/ml. I Pigment production on the 3rd day of incubation at 3 C. Symbols: +, Purple pigment produced; -, purple pigment not produced. Allelele numbers of the given mutants are Y175155, 3523, 3871, and Y112M13, respectively. Downloaded from on October 19, 218 by guest

6 VOL. 124, 1975 PURINE METABOLISM IN N. CRASSA 83 TABLE 3. Dry weights of ad3b and ad3a mutants, obtained at the times indicated after growing in adenine, adenine + histidine, and adenine + tryptophan media Dry wt (mg/52 ml of medium)a 72 h after inoculationb 12 h after inoculationc Strain Adenine (5 Adenine (5 Adenine (5 Adenine (5 Adenine pg/ml) + pg/ml) + Adenine pg/ml) + ug/ml) + (5 ug/ml) histidine tryptophan (5 plg/mi) histidine tryptophan (5 ug/ml) (5 pg/ml) (5 pg/ml) (5 pg/ml) ad3b (3523) ad3b (Y175M 155) ad3a (3871) ad3a (Y112M13) a 2 ml of spore suspension of each of the strains was added to 5 ml of medium. Dry weights obtained from the contents of duplicate flasks were averaged and presented above. 'At this time all strains were showing pigmentation in adenine medium but not in histidine medium. Traces of pigmentation were seen in some of the tryptophan-containing flasks. c At this time intense pigmentation was seen in adenine medium for all the strains, but small amounts of pigmentation were present in both histidine- and tryptophan-containing media. developed an amber color from a very early period after the media were inoculated. As reasoned earlier, if the purple pigment production can be equated with the de novo purine synthetic ability, the observed lag in pigmentation may be a consequence of inhibition of a step in purine synthesis. If histidine and tryptophan have any inhibitory effect at all on purine synthesis, one would expect that the high concentration of these amino acids would have an effect also on the growth of wild type. For this reason, the growth of wild type in minimal medium, histidine-supplemented medium, and tryptophan-supplemented medium was measured in terms of dry weight. For this experiment, a conidial suspension (4.7 x 17 conidia/ml) of wild-type strain was prepared and 2 ml of this suspension was inoculated into each of 4 ml of medium in 25-ml Erlenmeyer flasks. At specified times, total contents of duplicate flasks for each treatment were filtered and dry weights were averaged (Fig. 4). During 28 h of observations both histidine and tryptophan reduced the growth of wild type, but the reduction in growth in tryptophan-supplemented medium was far greater than that in histidine-supplemented medium. DISCUSSION The pool utilization studies presented here indicate that Neurospora has the same type of interrelationship between adenine and histidine pathways as described previously in bacteria and yeasts (1, 11, 13, 16, 19). Thus in adenine mutants of Neurospora blocked after AICAR formation (ad8 and ad4), some of the reserve 16/ ' 14_ 12_ C 1_ o 6_ TIME (HOURS) FIG. 4. Growth responses of wild-type Neurospora in minimal, histidine-supplemented, and tryptophansupplemented media. Erlenmeyer flasks containing 4 ml of each of the media were inoculated with 2 ml of spore suspension (4.7 x 17 spores/ml) and shaken at 3 C. Duplicate flasks of each medium were filtered for dry weights at the specified times and the weights were averaged. Symbols: () Minimal medium; (A) histidine-supplemented medium; () tryptophan-supplemented medium. nucleotides are apparently channeled into histidine synthesis. In the presence of histidine a sparing of these nucleotides occurs, which is reflected as an increase in the numbers of spores germinating. However, a comparable germination to that of ad9 and ad3b cannot be expected in these strains, since the bulk of conidial purine reserves is in the form of inosine and ad8 and ad4 mutants are inosine nonutilizers. The nature of nucleic acid degradation in ad8 and ad4 mutants in minimal medium is not known. Downloaded from on October 19, 218 by guest

7 84 PENDYALA AND WELLMAN Since only a net nucleic acid content was measured at any time, the sparing of nucleic acid breakdown in histidine medium in these strains may in fact be due to an increase in the nucleic acid synthesis in the presence of histidine. If both degradation and synthesis were taking place simultaneously, only the net outcome would be recorded. In view of the stimulated germination of conidia of ai8 and ad4 mutants in histidine-supplemented media and of the requirement of ribonucleic acid synthesis for germination in N. crassa (8), a simultaneous synthesis and degradation of some of the preexisting nucleic acid must in fact be taking place. The net amount of '4C-labeled nucleic acid recorded in these strains is higher in histidinesupplemented medium compared with that on minimal medium, indicating that more of the reserve pool was utilized for nucleic acid synthesis. It is interesting to note here that the leakage of labeled purines is also affected in strains ad8 and ad4 in the presence of histidine. From cross-feeding studies with adenine mutants, it was assessed that the leaking compound may be inosine or hypoxanthine and that the leakage occurs only when there is a carbon source in the medium (15). At this time it is not clear how histidine exerts an effect on a possible energydependent outward transport of either hypoxanthine or inosine. In addition to the specific cyclical interaction between adenine and histidine pathways described above, histidine seems to influence the de novo adenine nucleotide synthesis possibly by a different mechanism, since among several amino acids tested, not only histidine but also tryptophan was found to exert an inhibitory effect on purple pigment production in ad3b and ad3a mutants. If both of these amino acids affect the pigmentation by the same means, it may be due either to a direct inhibitory effect of these amino acids on the enzymes unique to adenine nucleotide biosynthetic pathway or to a lowering of the supply of PRPP. PRPP is a common precursor for histidine, tryptophan, and adenine pathways. The supply of PRPP is crucial for de novo purine synthesis (3, 7). Therefore, if the presence of histidine or tryptophan lowers the available PRPP for purine synthesis, the de novo purine synthesis would be affected. Although histidine and tryptophan are end products of divergent pathways, the synthesis of both these amino acids requires PRPP, and it therefore seems plausible that they might regulate the synthesis of PRPP by end-product inhibition of PRPP synthetase (EC ), the enzyme that catalyzes the formation of PRPP. J. BAennuoL. However, from the work on other organisms, there is no evidence that either of these amino acids in fact regulates the PRPP synthesis. R. L. Switzer (Fed. Proc. 26:56, 1967) reported that tryptophan, but not histidine, was inhibitory to Salmonella PRPP synthetase. Yet, his later studies on purified PRPP synthetase of Salmonella gave no evidence of inhibition of this enzyme either by histidine or tryptophan (18). In E. coli only a slight inhibition of PRPP synthetase by tryptophan was reported (2). Until further information is available on PRPP levels in Neurospora in the presence of histidine and tryptophan, the possibility remains that observed inhibition by histidine and tryptophan might be due either to a lowering of PRPP concentration or to a direct regulation of purine nucleotide biosynthetic enzymes. If the latter is found to be true, the adenine nucleotide pathway would be a part of the cross-pathway regulation described by Carsiotis and Jones (6) for Neurospora. ACKNOWLEDGMENTS This investigation was supported by National Research Council of Canada grant A1394. LITERATURE CITED 1. Armitt, S., and R. A. Woods Purine excreting mutants of Saccharomyces cerevisiae. 1. Isolation and genetic analysis. Genet. Res. 15: Atkinson, D. E., and L. Fall Adenosine triphosphate conservation in biosynthetic regulation. Escherichia coli phosphoribosyl pyrophosphate synthetase. J. Biol. Chem. 242: Bagnara, A. S., A. A. Letter, and J. F. Henderson Multiple mechanisms on regulation of purine biosynthesis de novo in intact tumor cells. Biochim. Biophys. Acta 374: Bernstein, H Imidazole compounds accumulated by purine mutants of Neurospora crassa. J. Gen. Microbiol. 25: Buchanan, J. M The enzymatic synthesis of the purine nucleotides. Harvey Lect. 54: Carsiotis,!A, and R. F. Jones Cross-pathway regulation: histidine-mediated control of histidine, tryptophan, and arginine biosynthetic enzymes in Neurospora crassa. J. Bacteriol. 119: Hartman, S. C Purines and pyrimidines, p In D. M. Greenberg (ed.), Metabolic pathways, vol. 4. Academic Press Inc., New York. 8. Hollomon, D. W Ribonucleic acid synthesis during fungal spore germination. J. Gen. Microbiol. 62: Jha, K. K An unlinked mutation affecting control of purine metabolism in a revertant of an ad 7 auxotroph of Neurospora crassa lacking phosphoribosyl pyrophosphate amidotransferase. Mol. Gen. Genet. 114: Jones, E. W., and B. Magasanik Phosphoribosyl-5- amino-4-imidazole carboxamide formyl transferase activity in the adenine-histidine auxotroph ad 3 of S. cerevisiae. Biochem. Biophys. Res. Commun. 29: Lomax, C. A., T. S. Gross, and R. A. Woods New Downloaded from on October 19, 218 by guest

8 VOL. 124, 1975 mutant types at the ade 3 locus of Saccharomyces cerevisiae. J. Bacteriol. 17: McElroy, W. D., and H. K. Mitchell Enzyme studies on a temperature sensitive mutant of Neurospora crassa. Fed. Proc. 5: Magasanik, B., and D. Karibian Purine nucleotide cycles and their metqbolic role. J. Biol. Chem. 235: Mitchell, H. K., and M. B. Houlahan Adenine requiring mutants of Neurospora crassa. Fed. Proc. 5: Pendyala, L., and A. M. Wellman Endogenous purine metabolism in the conidia of wild type and certain adenine mutants of Neurospora crassa. 1. The nature of reserve pools and pool utilization during adenine starvation. Biochim. Biophys. Acta 385: Sheldovsky, A. E., and B. Magasanik A defect in histidine biosynthesis causing an adenine deficiency. J. PURINE METABOLISM IN N. CRASSA 85 Biol. Chem. 237: Smirnov, M. N., V. N. Smirnov, E. I. Budowsky, S. G. Inge-Vechtomov, and N. G. Serebrjakov Red pigment of adenine deficient yeast Saccharomyces cerevisiae. Biochem. Biophys. Res. Commun. 27: Switzer, R. L., and D. S. Sogin Regulation and mechanism of PRPP synthetase. V. Inhibition by end products and regulation by adenosine diphosphate. J. Biol. Chem. 248: Whitehead, E. P La cinetique allosterique de l'inhibition du premier enzyme de la biosynthese de le histidine chez S. pombe. Bull. Soc. Chim. Biol. 49: Wyngaarden, J. B., and D. M. Ashton The regulation of phosphoribosyl pyrophosphate amidotransferase by purine ribonucleotides: a potential feedback control bf purine biosynthesis. J. Biol. Chem. 234: Downloaded from on October 19, 218 by guest