Synergistic and Product Induction of the Enzymes of Tryptophan Metabolism in Pseudomonas acidovorans

Tryptophan oxygenase (tryptophan 2,3-dioxygenase) activity increases immediately before the initiation of actinomycin D production by Streptomyces parvullus. We have attempted to discern whether this increase is due to a release from catabolite repression or to the synthesis of an inducer substance. The standard culture medium (glutamic acid-histidine-fructose medium) used in antibiotic production studies with S. parvullus contains L-glutamate as a major constituent. L-Glutamate is almost totally consumed before the onset of actinomycin D synthesis. The addition of 10 mM L-glutamate at this stage completely abolished actinomycin D production as well as tryptophan oxygenase synthesis. Fourteen amino acids were tested for a similar effect. Of these, L-glutamate and L-aspartate had the most dramatic effect on tryptophan oxygenase and ,8-galactosidase (fl-D-galactosidase), another inducible enzyme. Standard glutamic acidhistidine-fructose medium, preincubated for 23 h to remove L-glutamate, allowed the synthesis of actinomycin D and tryptophan oxygenase by cells at a stage of growth normally considered too early for antibiotic production. A chemically defined medium lacking L-glutamate and adjusted to pH 8.0 was designed to simulate the preincubation medium. The transfer of cells to this artificial preincubation medium resulted in the appearance of tryptophan oxygenase as early as 19 h before normal synthesis occurred, eliminating the possibility that an inducer molecule is synthesized and excreted during the preincubation period. The results of these studies suggest that the increase in tryptophan oxygenase activity before the onset of actinomycin D synthesis, as well as the synthesis of actinomycin D itself, is due to a release from L-glutamate catabolite repression.The antibiotic actinomycin D is a secondary metabolite produced by several Streptomyces species. Most of these species produce more than one form of actinomycin (5). Streptomyces parvullus is unusual in that it only synthesizes one class of actinomycin, class IV (12). Consequently, research into the biochemistry of antibiotic production in this organism is uncomplicated by the presence of multiple enzyme systems responsible for the production of more than one form of actinomycin.Tryptophan has been implicated as a precursor of the actinocin ring structure of actinomycin D (6, 7). In addition, Lingens and Vollprecht (9) have presented evidence that, in at least one streptomycete, NAD is also synthesized from tryptophan. Figure 1, therefore, presents the pathways of tryptophan metabolism as they are believed to occur in Streptomyces spp. (19).Tryptophan oxygenase (tryptophan 2,3-dioxygenase; L-tryptophan:oxygen 2,3-oxidoreductase; EC 1.13.11.11) is the first enzyme involved t Present address: in both the primary metabolic pathway to NAD and the secondary metabolic pathway in actinomycin D. Its activity was first demonstrated in S. parvullus by M. J. M. Hitchcock and E. Katz (unpublished data), who demonstrated its apparent induction during the onset of ac...

“…However, they were unable to experimentally induce tryptophan oxygenase with exogenously supplied tryptophan. This contrasts with the results obtained for other organisms (2,16).…”

contrasting

confidence: 99%

contrasting

confidence: 83%

Control of Actinomycin D Biosynthesis inStreptomyces parvullus: Regulation of Tryptophan Oxygenase Activity

Foster

Katz

1981

“…However, it is similar to the P. acidovorans pathway because 75 VOL. 121, 1975 on July 16, 2020 by guest http://jb.asm.org/ Downloaded from both use kynurenine as a gratuitous inducer of tryptophan oxygenase (14).…”

Section: Discussionmentioning

confidence: 99%

Tryptophan catabolism in Bacillus megaterium

Bouknight¹,

Sadoff²

1975

Bacillus megaterium grows in a medium containing L-tryptophan as the sole carbon, nitrogen, and energy source. Kynurenine, anthranilic acid, and catechol are metabolic intermediates, suggesting that this organism used the anthranilic acid pathway for tryptophan degradation. Cells that grow on L-tryptophan oxidize kynurenine, alanine, and anthranilic acid and the presence of tryptophan oxygenase (EC 1.13.1.12), kynureninase (EC 3.7.1.3), and catechol oxygenase (EC 1.13.1.1) in cell extracts provide additional evidence for the degradative pathway in B. megaterium. Tryptophan oxygenase is inhibited by sodium azide, potassium cyanide, and hydroxylamine, indicating that the enzyme has a functional heme group. D-Tryptophan is not a substrate for tryptophan oxygenase, and the D-isomer does not inhibit this enzyme. Formamidase (EC 3.5.1.9) and anthranilate hydroxylase are not detectable in extracts. Tryptophan catabolism is inducible in B megaterium and is subject to catabolite repression by glucose and glutamate. Arginine does not cause repression, and kynurenine induces both tryptophan oxygenase and kynureninase.

“…Tryptophan and hydrocortisone stabilize or activate and induce the formation of tryptophan pyrrolase in rat liver (14,28); tryptophan coordinately induces tryptophan pyrrolase, kynurenin formamidase, and kynureninase in X. pruni (4); N-formylkynurenin induces kynureninase in N. crassa (29). Similar enzymes are induced by tryptophan in bacterial species which do not use tryptophan as a precursor of nicotinic acid (25,27). Tryptophan pyrrolase activity has not been obtained in cell-free preparations of Neurospora or Saccharomyces (1).…”

mentioning

confidence: 99%

End-Product Regulation of the Tryptophan-Nicotinic Acid Pathway in Neurospora crassa

Lester

1971

The regulation of the tryptophan-nicotinic acid pathway in Neurospora crassa was examined with mutants (nic-2, nic-3) which require nicotinamide for growth. The accumulation of N-acetylkynurenin and 3-hydroxyanthranilic acid by these mutants served to estimate the level of function of the early reactions in the pathway. In still cultures, maximal accumulation occurred with media containing growth-limiting amounts of nicotinamide; the accumulation of intermediates was almost negligible with nicotinamide in excess. Only nicotinamide and closely related compounds which also supported the growth of these mutants inhibited the accumulation of intermediates. The site of inhibition was assessed to be between tryptophan and kynurenin (or N-acetylkynurenin). The synthesis of N-acetylkynurenin was examined in washed germinated conidia suspended in buffer; the level of N-acetylkynurenin-synthesizing activity was inversely related to the concentration of nicotinamide in the germination medium. The addition of large amounts of nicotinamide to suspensions of germinated conidia did not affect their N-acetylkynurenin-synthesizing activity. Formamidase activity, kynurenin-acetylating activity, and gross tryptophan metabolism in germinated conidia was not influenced by the concentration of nicotinamide in the germination medium. The results obtained indicate that the site of inhibition by nicotinamide is the first step in the pathway, the tryptophan pyrrolase reaction. The data are interpreted as nicotinamide or a product thereof, such as nicotinamide adenine dinucleotide, acting as a repressor of the formation of tryptophan pyrrolase in N. crassa.