Isolation and characterization of Neurospora crassa mutants impaired in feedback control of ornithine synthesis

Weiss, Richard L.; Lee, C A

doi:10.1128/jb.141.3.1305-1311.1980

Cited by 14 publications

(6 citation statements)

References 26 publications

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“…Another remarkable result, concerning the regulation of the first enzymes of the arginine pathway, has been reported by the team of R. L. Weiss. A series of ornithine-overproducing N. crassa mutants [33], were mapped to the N-terminus of N-acetylglutamate kinase and shown to bear F81L modifications. The data suggest that this single amino-acid modification of the kinase might result in the deregulation of the first two enzyme activities of the arginine pathway, leading to the hypothesis of a co-ordinated feedback control (R. L. Weiss, S. K. Chae, J. Chung, C. McKinstry, M. Karaman and G. Turner, University of California, Los Angeles, CA, USA, personal communication).…”

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TheN‐acetylglutamate synthase/N‐acetylglutamate kinase metabolon ofSaccharomyces cerevisiaeallows co‐ordinated feedback regulation of the first two steps in arginine biosynthesis

Pauwels

Abadjieva

Hilven

et al. 2003

European Journal of Biochemistry

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In Saccharomyces cerevisiae, which uses the nonlinear pathway of arginine biosynthesis, the first two enzymes, N-acetylglutamate synthase (NAGS) and N-acetylglutamate kinase (NAGK), are controlled by feedback inhibition. We have previously shown that NAGS and NAGK associate in a complex, essential to synthase activity and protein level The NAGKs of ascomycetes possess, in addition to the catalytic domain that is shared by all other NAGKs and whose structure has been determined, a C-terminal domain of unknown function and structure. Exploring the role of these two domains in the synthase/kinase interaction, we demonstrate that the ascomycete-specific domain is required to maintain synthase activity and protein level.Previous results had suggested a participation of the third enzyme of the pathway, N-acetylglutamylphosphate reductase, in the metabolon. Here, genetic analyses conducted in yeast at physiological level, or in a heterologous background, clearly demonstrate that the reductase is dispensable for synthase activity and protein level.Most importantly, we show that the arginine feedback regulation of the NAGS and NAGK enzymes is mutually interdependent. First, the kinase becomes less sensitive to arginine feedback inhibition in the absence of the synthase. Second, and as in Neurospora crassa, in a yeast kinase mutant resistant to arginine feedback inhibition, the synthase becomes feedback resistant concomitantly.We conclude that the NAGS/NAGK metabolon promotes the co-ordination of the catalytic activities and feedback regulation of the first two, flux controlling, enzymes of the arginine pathway.

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confidence: 99%

TheN‐acetylglutamate synthase/N‐acetylglutamate kinase metabolon ofSaccharomyces cerevisiaeallows co‐ordinated feedback regulation of the first two steps in arginine biosynthesis

Pauwels

Abadjieva

Hilven

et al. 2003

European Journal of Biochemistry

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“…The kinase therefore limits ornithine synthesis in conditions of arginine excess. Feedback-resistant mutants have been isolated (240). The kinase of these mutants has reduced sensitivity to arginine (50% inhibition at 14 mM), and the mutants, unlike the wild type, synthesize ornithine in vivo in the presence of arginine (I. Goodman and R. L. Weiss, submitted for publication).…”

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Compartmental and regulatory mechanisms in the arginine pathways of Neurospora crassa and Saccharomyces cerevisiae.

Davis¹

1986

Microbiological Reviews

136

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Arginine metabolism has been studied intensively in manly organisms, beginning in earnest with the discovery of the urea cycle in mammals (139). The genetic control of the arginine pathway was the earliest example of the one-gene, one-enzyme relationship in the biochemical genetics of Neiurosporca crassa (206). The elucidation of the pathway in Escherichia coli followed soon thereafter, the term "repression'' having been coined to describe the regulatory behavior of acetylornithinase (229). The discovery of carbamoyl phosphate in 1955 (131) led to many comparative studies of its metabolism (52, 130, 152). A profound knowledge of arginine metabolism in N. crassa and Sacclhiaonvces (ceev'isiae has developed in the last 25 years. The subject justifies a comparative review, because the two organisms solve similar metabolic problems in different ways. It is likely that the fundamental phenomena of compartmentation and regulation in the two organisms can be seen throughout the evolutionary tree, used variously according to the demands of particular lifestyles. The synthesis of arginine in fungi has three main components: the synthesis of ornithine, the synthesis of carbamoyl phosphate, and the conversion of these two compounds to arginine. The catabolic pathway consists of the hydrolysis of arginine to ornithine and urea, the breakdown of urea to ammonia and carbon dioxide, and the conversion of ornithine to glutamate. A theme to be pursued is how the anabolic and catabolic pathways can each proceed to the exclusion of the other, given ornithine as a common intermediate. A related theme is how carbamoyl phosphate and ornithine are confined to the arginine pathway in the face of enzymes that might divert them to other fates. Both of these matters involve the compartmentation of small molecules by intracellular membranes, which in S. (ereiisiae is supplemented by elaborate enzyme regulatory mechanisms. This review describes the enzymology, genetics, and localization of the arginine enzymes of S. cere\eisiae and N. crassa. This is followed by descriptions of vacuolar function, enzyme regulation, and the integration of relevant factors in adaptation to different environments. Metabolic maps (Fig. 1 and 2) and a gene-enzyme directory (Table 1) are given for N. crassa(and S. cerevisiae. Arginine metabolism in these organisms has been reviewed in more limited ways previously (1

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“…Citrulline synthesis in a feedback-resistant strain. The sup-3 mutation imparts feedback resistance to the intramitochondrial enzyme acetylglutamate kinase (37,41), an initial enzyme of ornithine synthesis (Fig. 1).…”

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confidence: 99%

Control of the ornithine cycle in Neurospora crassa by the mitochondrial membrane

Davis¹,

Ristow²

1983

J Bacteriol

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In Neurospora crassa, the mitochondrial membrane separates ornithine used in arginine biosynthesis from ornithine used in the arginine degradative pathway in the cytosol. Ornithine easily exchanges across the mitochondrial membrane under conditions appropriate for synthesis of the immediate biosynthetic product, citrulline. Neither of the two mitochondrial enzymes required for the ornithine-to-citrulline conversion is feedback inhibitable in vitro. Nevertheless, when arginine is added to cells and cytosolic ornithine increases as arginine degradation begins, the rate of citrulline synthesis drops immediately to about 20% of normal (B. J. Bowman and R. H. Davis, Bacteriol. 130:285-291, 1977). We have studied this phenomenon in citrulline-accumulating strains carrying the arg-1 mutation. Citrulline accumulation is blocked when arginine is added to an arg-1 strain but not to an arg-1 strain carrying a mutation conferring insensitivity of intramitochondrial ornithine synthesis to arginine. Thus, ornithine is evidently unable to enter mitochondria in normal (feedback-sensitive) cells. Other experiments show that cytosolic ornithine enters mitochondria readily except when arginine or other basic amino acids are present at high levels in the cells. We conclude that in N. crassa, the mitochondrial membrane has evolved as a secondary site of feedback inhibition in arginine synthesis and that this prevents a wasteful cycling of catabolic ornithine back through the anabolic pathway. This is compared to the quite different mechanism by which the yeast Saccharomyces cerevisiae prevents a futile ornithine cycle.

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Isolation and characterization of Neurospora crassa mutants impaired in feedback control of ornithine synthesis

Cited by 14 publications

References 26 publications

TheN‐acetylglutamate synthase/N‐acetylglutamate kinase metabolon ofSaccharomyces cerevisiaeallows co‐ordinated feedback regulation of the first two steps in arginine biosynthesis

TheN‐acetylglutamate synthase/N‐acetylglutamate kinase metabolon ofSaccharomyces cerevisiaeallows co‐ordinated feedback regulation of the first two steps in arginine biosynthesis

Compartmental and regulatory mechanisms in the arginine pathways of Neurospora crassa and Saccharomyces cerevisiae.

Control of the ornithine cycle in Neurospora crassa by the mitochondrial membrane

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