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Query: EC:3.5.1.4 (deaminase)
 5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe the regulatory properties of two strains carrying either the ilvA624 or the ilvA625 mutations, located in the structural gene for threonine deaminase. Crude extracts of both these strains possess a threonine deaminase activity migrating on polyacrylamide gels, differently from the wild type enzyme. Growth studies demonstrate that these mutations do not cause a limitation of isoleucine biosynthesis, suggesting normal catalytic activity of deaminase. A regulatory consequence of the ilvA624 allele is a derepression of the isoleucine-valine biosynthetic enzymes, which is recessive to an ilvA+ allele. The ilvA625 mutation causes a derepression which is dominant in an ilvA625/ILVA+ diploid. We interpret these data assuming that threonine deaminase, previously shown to be an autogenous regulator of the ilv genes, lacks a repressor function in the ilvA624 mutant, while in the ilvA625 mutant it is a better activator than wild type threonine deaminase. The data are discussed in terms of a model requiring that threonine deaminase, or a precursor of it, is in equilibrium between two forms, one being an activator of gene expression and the other being a repressor.
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PMID:Dual autogenous regulatory role of threonine deaminase in Escherichia coli K-12. 34 81

Kinetic analysis of the biosynthetic threonine deaminase, EC 4.2.1.16, from Samonella typhimurium yields hyperbolic substrate saturation curves in the absence of, and higher order substrate saturation curves in the presence of, L-isoleucine. L-Valine reverses this effect of L-isoleucine by restoring the hyperbolic substrate saturation curves. The inhibition of enzyme activity and the reversal of valine stimulation is a function of a second order concentration of L-isoleucine, whereas antagonism of inhibition is a function of first order concentration of valine. The antagonistic effects on enzyme activity of L-isoleucine and of L-valine appear as competitive in diagnostic plots. Threonine deaminase possesses two L-isoleucine binding sites (Kd equals 3.6 muM) and one L-valine binding site (Kd equals 26 muM); the binding of these ligands appear competitive. Exclusion of L-valine requires the binding of 2 molecules of L-isoleucine whereas binding of a single L-valine molecule prevents the binding of 2 L-isoleucine molecules. Cooperative binding of L-isoleucine is not observed under any of the conditions tested. Two cases, expressed in terms of modified Adair equations and based upon the assumption that L-threonine also serves as an activator ligand which binds to the L-valine site, are presented. Case I states that liganding of the activator sites must percede substrate-binding at the active site, and Case II states that the activator site liganding is required solely for reactivation of the L-isoleucine-inhibited enzyme. Analysis of kinetic data by a curve-fitting process suggests that Case II described the relationship between the activator site and the L-isoleucine sites. An enzymatically inactive derivative of threonine deaminase, prepared by reduction with borohydride, binds isoleucine and valine in a manner similar to native holoenzyme. Binding of L-threonine and L-valine to the derivatized enzyme is competitive. The Kd for threonine binding is 3 mM, which is in excellent agreement with the Kd determined by the curve fitting process. It is concluded that the modulation of threonine deaminase activity is wrought by interaction between inhibitor sites and an activator site rather than inhibitor and active sites and that induced transitions rather than concerted transitions more adequately describe the underlying regulatory principle.
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PMID:Threonine deaminase from Salmonella typhimurium. Relationship between regulatory sites. 108 62

In confirmation of the findings of Gaitonde et al. (1974), a decrease in the brain concentration of threonine and serine, and an increase in glycine, were observed in rats maintained on a thiamin-deficient diet. Similar changes were found in the blood, and the concentration of several other amino acids in the blood decreased significantly. There was a correlation between the concentrations of threonine, serine, aspartate and asparagine in the brain and blood. In experiments in which [U-14C]threonine was injected into rats most of the radioactivity in the brain and blood of control rats was, as expected, in threonine in the acid soluble metabolites. In contrast, a considerable proportion of radioactivity was also found in other amino acids, namely glutamate, glutamine, aspartate, gamma-aminobutyrate and alanine, in the brain of thiamin-deficient rats. [U-14C]Threonine was also converted into 14C-labelled lactate and glucose, but the extent of this conversion was severalfold higher in thiamin-deficient than in control rats. This finding gave evidence of the stimulation in thiamin-deficient rats of the catabolism of [U-14C]threonine to [14C]lactate by the aminoacetone pathway catalysed by threonine dehydrogenase, and into succinate via propionate by the alpha-oxobutyrate pathway catalysed by threonine dehydratase (deaminase). The measurement of specific radioactivities of glutamate, aspartate and glutamine after injection of [U-14C]threonine, indicated a stimulation of the activities of threonine dehydrogenase and threonine dehydratase (deaminase) in the brain of thiamin-deficient rats. The specific radioactivities of glutamate, asparatate and glutamine int he brain were consistent with an alteration in the metabolism of threonine, mainly in the 'large' compartment of the brain of thiamin-deficient rats. The measurement of relative specific radioactivity of proteins after injection of [U-14C]threonine indicated a marked decrease in the synthesis of proteins, mainly in the liver of thiamin-deficient rats.
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PMID:Conversion of [U-14C]threonine into 14C-labelled amino acids in the brain of thiamin-deficient rats. 118 Sep 21

The ilvA gene of Pseudomonas cepacia was expressed poorly in Escherichia coli. Insertion of IS2 upstream of the cloned gene dramatically increased its transcription, resulting in an 85-fold increase in threonine dehydratase (deaminase) specific activity. The results confirm earlier reports that IS2 promotes efficient expression of foreign genes in E. coli.
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PMID:IS2 activates the ilvA gene of Pseudomonas cepacia in Escherichia coli. 303 Oct 23

l-Threonine deaminase (l-threonine dehydratase [deaminating], EC 4.2.2.16) has been shown to be involved in the regulation of three of the enzymes of isoleucine-valine biosynthesis in yeast. Mutations affecting the affinity of the enzyme for isoleucine also affected the repression of acetohydroxyacid synthase, dihydroxyacid dehydrase, and reductoisomerase. The data indicate that isoleucine must be bound for effective repression of these enzymes to take place. In a strain with a nonsense mutation midway in liv 1, the gene for threonine deaminase, starvation for isoleucine or valine did not lead to derepression of the three enzymes; starvation for leucine did. The effect of the nonsense mutation is recessive; it is tentatively concluded, therefore, that intact threonine deaminase is required for derepression by two of the effectors for multivalent repression, but not by the third. A model is presented which proposes that a regulatory species of leu tRNA(leu) is the key intermediate for repression and that threonine deaminase is a positive element, regulating the available pool of charged leu tRNA by binding it.
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PMID:Involvement of threonine deaminase in repression of the isoleucine-valine and leucine pathways in Saccharomyces cerevisiae. 457 Jul 83

Threonine deaminase (l-threonine dehydratase EC 4.2.1.16) has been partially purified from a new extreme thermophilic bacterium, Thermus X-1, which is similar to T. aquaticus YT-1. The threonine deaminase of strain X-1 has a maximal rate of reaction at 85 to 90 C and is more thermostable than the threonine deaminase from mesophilic bacteria. The enzyme has an apparent molecular weight of 100,000 to 115,000, a K(m) for l-threonine of 14 mM, a pH optimum of 8.0, and like other threonine deaminases also catalyzes the deamination of serine. However the Thermus X-1 threonine deaminase does not show a strong feedback inhibition by isoleucine. It is suggested that the regulation of the biosynthesis of isoleucine in this extreme theromophile may resemble that reported in Rodospirillum rubrum.
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PMID:Purification and properties of threonine deaminase from the X-1 isolate of the genus Thermus. 470 89

Saccharomyces cerevisiae mutants lacking the anabolic L-threonine deaminase, the ilv1- mutants, have been found to exhibit a normal ability to grow, without auxotrophy towards isoleucine, on L-threonine of L-serine as only nitrogen nutrient. Starting from a strain carrying a ilv1- mutation, a new mutation affecting the ability to utilize L-threonine as nitrogen source was selected. This mutation, which also impairs the ability to utilize L-serine, has been denominated cha-, for 'catabolism of hydroxyamino acids' and was found to result in the lack of a catabolic L-serine (L-threonine) deaminase. This enzyme which, unlike the anabolic threonine deaminase, is more active towards serine than towards threonine, differs from the latter enzyme by a number of biochemical and regulatory properties. Whereas the anabolic enzyme is an allosteric enzyme sensitive to feedback inhibition by isoleucine, the catabolic enzyme exhibits Michaelian kinetics: no control of its activity has been detected. Its synthesis is induced by L-serine and L-threonine. These two enzymes, which thus can be easily differentiated by means of their regulations, display a limited ability to compensate for one another's absence and appear to play clearly distinct roles under normal physiological conditions.
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PMID:Occurrence of a catabolic L-serine (L-threonine) deaminase in Saccharomyces cerevisiae. 704 46

L-2-Aminobutyric acid was synthesised in a transamination reaction from L-threonine and L-aspartic acid as substrates in a whole cell biotransformation using recombinant Escherichia coli K12. The cells contained the cloned genes tyrB, ilvA and alsS which respectively encode tyrosine aminotransferase of E. coli, threonine deaminase of E. coli and alpha-acetolactate synthase of B. subtilis 168. The 2-aminobutyric acid was produced by the action of the aminotransferase on 2-ketobutyrate and L-aspartate. The 2-ketobutyrate is generated in situ from L-threonine by the action of the deaminase, and the pyruvate by-product is eliminated by the acetolactate synthase. The concerted action of the three enzymes offers significant yield and purity advantages over the process using the transaminase alone with an eight to tenfold increase in the ratio of product to the major impurity.
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PMID:Engineering of a novel biochemical pathway for the biosynthesis of L-2-aminobutyric acid in Escherichia coli K12. 1057 28

Kerwar, Suresh S. (Oregon State University, Corvallis), Vernon, H. Cheldelin, and L. W. Parks. Valine-isoleucine metabolism in Acetobacter suboxydans and the inhibition of growth by valine. J. Bacteriol. 88:179-186. 1964.-Extracts of Acetobacter suboxydans can synthesize valine and isoleucine via acetolactate and acetohydroxybutyrate, respectively. The amounts of these amino acids synthesized from different intermediates were determined. The pathways appear to be identical to those described for yeast, Neurospora, and Escherichia coli. When exogenous valine was added to a synthetic growth medium inoculated with A. suboxydans, no growth of the culture was observed. The inhibitory effect of valine was reversed by the addition of isoleucine. The site and mechanism of valine inhibition were investigated. Threonine deaminase was inhibited by valine and isoleucine but not by leucine. Repression of the deaminase by isoleucine but not by valine was indicated. The data reported in this paper suggest that valine prevented growth of the organism through false feedback inhibition of threonine deaminase, thereby limiting isoleucine biosynthesis.
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PMID:VALINE-ISOLEUCINE METABOLISM IN ACETOBACTER SUBOXYDANS AND THE INHIBITION OF GROWTH BY VALINE. 1419 85

We fused four mutant omr1 alleles, encoding feedback-insensitive forms of Arabidopsis thaliana biosynthetic threonine dehydratase/deaminase (TD), to the CaMV 35S promoter and transformed these constructs into A. thaliana Columbia wild type plants. The mutant TD forms consisted of our previously isolated double mutant, omr1-1 , and three new site-directed mutants, omr1-5 , omr1-7 , and omr1-8 with single point mutations. We employed site-directed mutagenesis to assay the effects of amino acid substitutions in separate regulatory regions within the carboxy-terminal (C-term) allosteric end. TD assays and growth resistance to the isoleucine (Ile) toxic analog -O-methylthreonine (OMT) confirmed the desensitization to feedback inhibition and the viability of these mutant omr1 alleles as selectable markers, respectively. Two of the site-directed mutants, omr1-5 and omr1-7 , appeared to influence one of the two separate Ile-binding sites and had a notable 13-fold and 15-fold increase in free Ile, respectively. The omr1-8 appeared to influence the other Ile-binding site and resulted in a 2-fold increase in free Ile. The transgenic omr1-1 double mutant affecting both Ile-binding sites, however, displayed a 106-fold increase in free Ile revealing a profound synergistic interplay between these separate Ile-binding sites. While all of the four omr1 alleles conferred resistance to elevated concentrations of OMT, the progeny of omr1-1 initial transformants exhibited a bushy phenotype at the rosette stage. On the other hand, progeny of transformants omr1-5 , omr1-7 , and omr1-8 had a normal phenotype, undistinguishable from wild type. Therefore, alleles omr1-5 , omr1-7 , and omr1-8 , proved to be ideal as environmentally-friendly, dominant, selectable markers for plant transformation.
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PMID:A site-directed mutagenesis interrogation of the carboxy-terminal end of Arabidopsis thaliana threonine dehydratase/deaminase reveals a synergistic interaction between two effector-binding sites and contributes to the development of a novel selectable marker. 1560 69


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