The degradation of L-histidine, imidazolyl-L-lactate and imidazolylpropionate by Pseudomonas testosteroni.
(17/33)
1. Imidazol-5-ylpropionate and imidazol-5-yl-lactate are degraded by Pseudomonas testosteroni via inducible pathways. 2. Growth on either compound as the sole source of carbon results in the induction of the enzymes for histidine catabolism. 3. The pathway of histidine degradation in this organism, a non-fluorescent Pseudomonad, is shown to be the same as that operating in Pseudomonas fluorescens and Pseudomonas putida. It consists of the successive formation of urocanate, imidazol-4-on-5-ylpropionate, N-formimino-l-glutamate, N-formyl-l-glutamate and glutamate. 4. Whole cells of P. testosteroni accumulate urocanate in the reaction mixture when incubated with imidazolylpropionate, but only after an adaptive lag period which is removed by previous growth on imidazolylpropionate as the source of carbon. 5. Imidazolyl-lactate is oxidized to imidazolylpyruvate, which then gives rise to histidine by specific transamination with l-glutamate. 6. Cells grown on histidine, urocanate or imidazolylpropionate are also able to degrade imidazolyllactate. 7. Mutants lacking urocanase are unable to grow on imidazolylpropionate, imidazolyl-lactate, histidine or urocanate. One with impaired histidase activity cannot utilize histidine or imidazolyl-lactate, but grows normally on imidazolylpropionate or urocanate. A mutant unable to grow on imidazolylpropionate is indistinguishable from the wild-type with respect to growth on histidine, imidazolyl-lactate or urocanate. 8. Thus it is established that imidazolyl-lactate is metabolized via histidine whereas imidazolylpropionate enters the histidine degradation pathway after conversion into urocanate. (+info)
The control of the enzymes degrading histidine and related imidazolyl derivates in Pseudomonas testosteroni.
(18/33)
1. The induction of the enzymes for the degradation of l-histidine, imidazolylpropionate and imidazolyl-l-lactate in Pseudomonas testosteroni was investigated. 2. The activities of histidine ammonia-lyase, histidine-2-oxoglutarate aminotransferase and urocanase are consistent with these enzymes being subject to co-ordinate control under most growth conditions. However, a further regulatory mechanism may be superimposed for histidase alone under conditions where degradation of histidine must take place for growth to occur. 3. Experiments with a urocanase(-) mutant show that urocanate is an inducer for the enzymes given above and also for N-formiminoglutamate hydrolyase and N-formylglutamate hydrolase. 4. N-Formiminoglutamate hydrolase and N-formylglutamate hydrolase are also induced by their substrates, and it is suggested that these two enzymes may be different gene products from those expressed in the presence of urocanate. 5. Induction of the enzyme system for the oxidation of imidazolylpropionate is dependent on exposure of cells to this compound. (+info)
Enzymatic production of urocanic acid by Achromobacter liquidum.
(19/33)
To develop an efficient method for the production of urocanic acid, optimal conditions for the production of microbial L-histidine ammonia lyase and for the conversion of L-histidine to urocanic acid by this enzyme were studied. A number of microorganisms were screened to test their ability to form and accumulate urocanic acid from L-histidine. Achromobacter liquidum was selected as the best organism. With this organism, enzyme activity as high as 2.0 units/ml could be produced by a shaking culture at 30 C in a medium containing glucose, urea, potassium phosphate, L-histidine, yeast extract, peptone, and inorganic salts. Appropriate addition of a surface-active agent to the reaction mixture shortened the time required for the conversion. A large amount of L-histidine was converted stoichiometrically to urocanic acid in 48 h at 40 C. Accumulated urocanic acid was readily isolated in pure form by ordinary procedures with isoelectric precipitation. Yields of isolated urocanic acid of over 92% from L-histidine were easily attainable. When the culture of Achromobacter liquidum was added to DL-histidine, D-histidine and urocanic acid were simultaneously obtained in high yields. (+info)
Effect of temperature on histidine ammonia-lyase from a psychrophile, Pseudomonas putida.
(20/33)
Pseudomonas putida was able to grow at 0 C in a complex medium containing l-histidine and to synthesize histidine ammonia-lyase and urocanase. The activity of the former enzyme was assessed between -10 and 60 C in cells and in cell extracts. Activity was maximal from 20 to 35 C. Below 20 C, activity decreased with temperature but, significantly, the enzyme exhibited 30% of its maximal activity at 1.5 C. The temperature response was similar in both intact cells and cell extracts, which indicated that the cell membrane did not significantly limit the entry of histidine at low temperature. Above and below the maximal temperature range, the reduced activity was not caused by irreversible inactivation, as shown by preincubation experiments. Also, when the temperature was rapidly changed from 60 to 30 C during an assay, the reaction rate increased abruptly to the full 30 C activity without a lag. This demonstrated the rapid reversibility of inactivation. The apparent Michaelis constant increased with temperature. As the substrate concentration was decreased, the enzyme activity became less dependent on temperature. The efficiency of substrate entry and catalysis near 0 C are factors in the ability of this facultative psychrophile to grow in a histidine medium at 0 C. (+info)
Involvement of threonine dehydratase in biosynthesis of the alpha-ketobutyrate prosthetic group of urocanase.
(21/33)
Seventeen mutants of Pseudomonas putida that were unable to grow on threonine as nitrogen source owing to a lack of threonine dehydratase were isolated, and all were found to be unable to synthesize active urocanase. Spontaneous revertants selected for urocanase production concomitantly regained threonine dehydratase. Mutants that were unable to utilize urocanate as carbon source were also isolated, and these were defective in urocanase formation but were normal in threonine dehydratase levels. Since alpha-ketobutyrate is the prosthetic group for urocanase, these results are consistent with the proposal that threonine dehydratase is necessary for urocanase prosthetic group biosynthesis. However, the lack of urocanase activity in threonine dehydratase-negative mutants was shown not to be the result of reduced levels of endogenous free alpha-ketobutyrate, nor to the participation of threonine dehydratase in the initiation of urocanase biosynthesis through the conversion of threonyl-tRNA(Thr) to alpha-ketobutyryl-tRNA(Thr). Other alternatives for the participation of threonine dehydratase in urocanase biosynthesis are discussed. (+info)
Isolation of a trans-dominant histidase-negative mutant of Salmonella typhimurium.
(22/33)
A mutation of Salmonella typhimurium was obtained that results in the failure of cells to synthesize the enzyme l-histidine ammonia-lyase (histidase). The mutation mapped within the hutH gene and in merodiploid strains was dominant over the wild-type allele. Extracts from cells bearing the trans-dominant histidase-negative allele were shown to contain material that reacts immunologically with antiserum against purified wild-type histidase. It is proposed that the trans-dominant allele results in the synthesis of defective histidase subunits that can combine with, and partially inactivate, wild-type histidase subunits. This subunit mixing presumably does occur, as the enzyme synthesized in a hybrid merodiploid strain is abnormally heat sensitive. (+info)
Mechanistic study of the urocanase reaction using deuterated substrates and 1H-NMR spectroscopy.
(23/33)
1. Samples of (alpha-2H1, 5-2H1) and (alpha-2H1, beta-2H1) urocanic acid were prepared by a combination of chemical and enzymic methods. 2. The enzymic conversion of unlabelled urocanate was followed by 1H-NMR spectroscopy at 500 MHz in deuterium oxide. It was found (a) that urocanase promotes the exchange of the 5-hydrogen atom of the substrate faster than it catalyses the overall reaction, (b) that the product is an equilibrium mixture of racemic beta-(5-oxoimidazol-4-yl)propionate and beta-(5-hydroxyimidazol-4-yl)propionate and (c) that beta-(5-oxoimidazol-4-yl)-propionate is spontaneously hydrolysed under physiological conditions to N-formylisoglutamine. The rate of this hydrolysis is considerably diminished at +8 degrees C. 3. It was shown by ultraviolet and 1H-NMR spectroscopic measurements that beta-(5-hydroxyimidazol-4-yl)-propionate (gamma max approximately equal to 234 nm) exists in protonated from at low pH (less than 1) whereas pH (approximately equal to 7.5) it exists in equilibrium with beta-(5-oxoimidazol-4-yl)propionate (gamma max approximately equal to 269 nm). 4. (alpha-2H1, beta-2H1)Urocanate was reacted with urocanase in deuterium oxide. 1H-NMR spectroscopy at 500 MHz showed a slight incorporation of protium into the side-chain of the product. The incorporated protium corresponded roughly to the protium contamination of the solvent and was equally distributed between the alpha and beta positions. No transfer of the 5-hydrogen atom to the side-chain was detected. 5. Kinetic deuterium isotope effects of between 2 and 3 were measured when the urocanase reaction was conducted in deuterium oxide at different p2H values. 6. Implications of these findings for the mechanism of action of urocanase are discussed. (+info)
Effect of zinc deficiency on histidine metabolism in rats.
(24/33)
The effects of feeding a diet deficient in zinc (Zn) to male rats on histidine metabolism were studied. Results showed that significantly higher percentages of DL-histidine-carboxyl-14C and L-histidine-2-(ring)-14C were oxidized by Zn-deficient rats. The incorporation of L-histidine-2-(ring)-14C into the proteins of skin, muscle, and kidney were significantly reduced in Zn-deficient rats as compared to Zn-supplemented rats. Conversely, the radioactivity of liver protein of Zn-deficient rats was significantly increased. Zn deficiency increased the activities of liver histidase and urocanase but had no effect on the activity of liver histidine-pyruvate transaminase. The increases of enzymatic activities were not due to food intake and can be prevented upon Zn repletion. The liver of Zn-deficient rats contained normal amount of histidine but a reduced quantity of histamine. The results on urinary excretion indicated that Zn-deficient rats discharged the same amounts of one-methyl and three-methyl histidine as Zn-supplemented pair-fed rats. Overall findings support in principle the concept that Zn deficiency results in disturbances of protein metabolism and also indicate that Zn is an important factor in regulating histidine metabolism through the urocanic acid pathway. (+info)