Factors controlling the expressed activity of histidine ammonia-lyase in the epidermis and the resulting accumulation of urocanic acid.

The synthesis of urocanic acid by histidine ammonia-lyase in guinea-pig epidermis was investigated in various ways. 1. In epidermal homogenates the enzyme obeys Michaelis-Menten kinetics and shows marked dependence of its activity of pH, such that below pH 6 it is inactive. 2. Part-thickness skin samples cultured with radioactive histidine do not accumulate detectable radioactive urocanic acid during 3 days in culture. 3. Very little histidine ammonia-lyase activity can be detected in the living cells of the epidermis. The enzyme is almost completely restricted to the dead cells of the stratum corneum. 4. Radioactive histidine injected into living animals does not result immediately in the accumulation of radioactive urocanic acid. By 3 days after the injection, however, both radioactive urocanic acid and histidine appear, apparently at the expense of radioactive proteins, 5. In isolated stratum corneum, the residual histidine can be converted into urocanic acid by the histidine ammonia-lyase in the tissue only if the natural acidity of the tissue is neutralized. It is concluded from these observations that the biosynthesis of urocanic acid occurs naturally only in the stratum corneum, which contains only dead cells. The amount of urocanic acid accumulated is limited by the availability of free histidine produced by proteolysis of residual protein in these cells and by the penetration into the stratum corneum of the 'acid mantle' of the skin.     

Effect of dietary histidine content on the change in content of skin urocanic acid isomers in hairless mice irradiated with ultraviolet B

Hairless mice were fed with a 10% amino acid mixture diet (control diet, 0.42% histidine content), the control diet without histidine (histidine-free diet), or the control diet rich in histidine (histidine-rich diet; histidine content, 4.2%) for 32 days. They were irradiated with UV light of 312 nm for 30 min, and skin samples were periodically taken for measuring the urocanic acid isomers. Total urocanic acid isomers were decreased by UV irradiation in all the three groups, the recovery being the fastest in the histidine-rich group. The percentage increase in cis-urocanic acid/total urocanic acid was quickly increased by UVB irradiation. The recovery of the ratio was slightly higher in the histidine-rich group, although the total urocanic acid level was higher in the histidine-rich group than in the others. Therefore, the absolute cis-urocanic acid content in the skin was almost the same among the three groups. These results indicate that the increased histidine intake strengthened UVB protection without any decrease in immune suppression.     

Quantitative analysis of histidine and cis and trans isomers of urocanic acid by high-performance liquid chromatography: a new assay method and its application

To elucidate the factors involved in dry skin and the skin damage caused by UV light, it is necessary to analyze small amounts of stratum corneum to determine amino acid contents. A new assay method for this purpose is described. Dabsylated amino acids including histidine and the cis and trans isomers of urocanic acid were analyzed quantitatively by high-performance liquid chromatography (HPLC), using a reversed-phase column. Histidine and the isomers of urocanic acid were separated from 36 other amino acids thought to be present in the extract of stratum corneum. In the presence of the 36 amino acids, standard calibration curves were obtained from 0.25 to 2.5 pmol/μl, for histidine and for both isomers of urocanic acid. The coefficients of variation for the reproducibility of the analysis at 1.0 pmol/μl were 3.8%, 2.9% and 2.5% for the cis and trans isomers of urocanic acid and for histidine, respectively. Amounts of 2 to 50 pmol of cis and trans isomers of urocanic acid and histidine in the stratum corneum were detected. The ratio of the cis to the trans isomer of urocanic acid in sunburned stratum corneum was more than three times that in normal stratum corneum. This method appears to be useful for the determination of small amounts of histidine and of the cis and trans isomers of urocanic acid in the stratum corneum.     

Reversible reaction via a carbanion intermediate in the elimination of ammonia from l-histidine catalysed by histidine ammonia-lyase

l-[5′-²H₂]Histidine was used as a substrate to investigate the enzymatic reaction mechanism with histidine ammonia-lyase from Pseudomonas fluorescens. The study was performed to determine the exchange rate of deuterium at C-5′ of the imidazole ring with solvent hydrogen relative to the net urocanic acid production. The extent of hydrogen exchange at C-5′ of histidine or urocanic acid was measured by gas chromatography—mass spectrometry—selected ion monitoring, monitoring the molecular ion intensities of the respective gas chromatographic derivatives, at m/z 380 and 379 for histidine and at m/z 267 and 266 for urocanic acid. The observed hydrogen exchange at C-5′ suggested a reversible mechanism via a carbanion intermediate in the reaction with histidine ammonia-lyase.     

Reversible stepwise mechanism involving a carbanion intermediate in the elimination of ammonia from L-histidine catalyzed by histidine ammonia-lyase.

L-Histidine labeled with deuterium at the C-5' position of the imidazole ring, L-[5'-2H]histidine (His-5'-D), was used as a probe for investigating a stepwise reversible mechanism via a carbanion intermediate in the elimination of ammonia catalyzed by histidine ammonia-lyase (EC 4.3.1.3). The labeled L-histidine (His-5'-D) (2.45 mM) was incubated with histidine ammonia-lyase (200 units) from Pseudomonas fluorescens at pH 7.0 or 9.0 at 25.0 degrees C for 24 h. The time course of the reaction was examined to determine the rates of enzyme-catalyzed hydrogen exchange at C-5' of L-histidine and urocanic acid. The finding of the enzyme-catalyzed hydrogen exchange at C-5' of both L-histidine and urocanic acid in the presence of L-histidine provided a rational explanation for a stepwise reversible mechanism via a carbanion intermediate in the elimination reaction. The rate of increase in the concentration of urocanic acid exchanged with hydrogen (UA-5'-H) did not depend on the formation rate of urocanic acid and UA-5'-H was continuously formed at a constant rate (25.6 microM/h) even after the completion of urocanic acid formation. These observations suggested the presence of the reversible reaction of urocanic acid and a carbanion intermediate. Since there was only a minor contribution for the formation of UA-5'-H from L-histidine exchanged with solvent hydrogen (His-5'-H), the main pathway in the enzymatic reaction of His-5'-D must be the formation of UA-5'-D via a carbanion intermediate (carbanion-D). Regeneration of the carbanion-D from UA-5'-D by its reverse reaction and subsequent hydrogen incorporation at C-5' would contribute to a large extent for the formation of UA-5'-H. The stability of carbanion was also demonstrated to be approximately three times higher at pH 7.0 than at pH 9.0.     

Regulation of hut enzymes and intracellular protease activities in Vibrio alginolyticus hut mutants

The production of alkaline protease, collagenase and histidine utilization (Hut) enzymes by Vibrio alginolyticus wild-type, hutH1 and hutU1 strains was investigated. Alkaline protease synthesis was stimulated by histidine and urocanic acid in the wild-type and hutU1 strains. The hutH1 mutant alkaline protease production was stimulated by urocanic acid and not by histidine. The Hut enzymes in the wild-type strain were coordinately induced by histidine. Urocanase and formimino-hydrolase were induced by histidine in the hutH1 mutant which lacked histidase and was not able to convert histidine to urocanic acid. Collagenase production in peptone medium was inhibited in the hut mutants. It is concluded that in V. alginolyticus urocanic acid regulates alkaline protease synthesis but that the Hut enzymes are induced by histidine. The involvement of the Hut genetic system in the regulation of alkaline protease and collagenase synthesis is discussed.     

Disruption of histidine catabolism in NEUT2 mice

Homozygous NEUT2 mice lack cytosolic 10-formyltetrahydrofolate dehydrogenase (FDH; Champion et al. (1994) Proc. Natl. Acad. Sci. USA 91, 11,338-11,342) and as a consequence should be unable to oxidize carbon 2 of l-histidine to CO2 via 10-formyltetrahydrofolate in liver cytosol. There was essentially no oxidation of l-[2-14C]histidine to 14CO2 in homozygous NEUT2 mice, but 52% of the [2-14C]l-histidine dose was recovered in the urine within 24 h. Analysis of urine samples for [14C]formiminoglutamate, the expected excretion product, was negative; however, [14C]urocanic acid was detected. Investigation of histidine catabolism via the folate-dependent deamination pathway revealed no detectable urocanase activity in homozygous NEUT2 mice, while heterozygous NEUT2 mice had 50% urocanase activity compared to normal mice. Histidase and formiminotransferase-cyclodeaminase, also on the histidine deamination pathway, had similar specific activities in normal and NEUT2 mice. Histidine-pyruvate aminotransferase, the first enzyme of the alternate histidine transamination catabolic pathway did not appear to be affected by the loss of urocanase. Based on the excretion of urocanic acid it is estimated that NEUT2 mice catabolize approximately 40 micromol/day via the deamination pathway. The loss of urocanase activity in homozygous NEUT2 mice may allow these mice to survive the disruption in folate metabolism by sparing the liver cytosolic tetrahydrofolate pool.     

The enzymatic conversion of L-histidine to urocanic acid by whole cells of Micrococcus luteus immobilized on carbodiimide activated carboxymethylcellulose.

Whole cells of Micrococcus luteus (formerly Sarcina lutea ATCC 9341) have been covalently linked to a carboxymethylcellulose support system, with the retention of histidine ammonia-lyase activity. The dependence of the rate of urocanic acid formation on pH, temperature, and added surfactant concentration was similar for the free and the immobilized cells. The immobilization procedure used is based on the carbodiimide activation of carboxymethylcellulose and has been optimized for the histidine ammonia-lyase activity of the immobilized cells on a given weight of cellulose. In a column reactor at 23 degrees C and superficial velocity of 0.044 cm/min, 5 g of cellulose with bound cells gave a 35% conversion of an L-histidine solution (0.25M, pH 9.0) to urocanic acid for 16 days of continuous operation. The scope of this carbodiimide assisted immobilization procedure has been investigated for a series of microorganisms and a variety of carboxylate functionalized supports.     

Urocanic Acid Production by Sporogenous Bacteria

As the results of a screening of several type cultures of bacteria, Bacillus subtilis var. thermophilus was revealed to be the most powerful strain for urocanic acid production. The accumulation of urocanic acid by this bacteria is caused by deamination of L-histidine, and is particularly accelerated in the presence of a component (X-factor) in meat extract. In the decomposition of urocanic acid the optimal pH of urocanase activity is markedly inhibited by the deviation of pH of the culture medium. The histidase of this bacteria is supposed to be a new exo-type enzyme.     

Effect of starvation on free histidine and amino acids in white muscle of milkfish Chanos chanos

Milkfish (Chanos chanos) decreased their body weight from 47 to 28 g over the 60-day period of starvation. Starvation also resulted in the reduction of muscle lipid and protein, and hepatosomatic index. The predominant free amino acid (FAA) in white muscle of milkfish was histidine, followed by taurine and glycine. In the first 25 days of starvation, no significant change in histidine was found. After 40 days of starvation, however, the histidine concentration was significantly decreased by 46%, and remained unchanged thereafter. As compared to control group fish, the 60-day-starved fish possessed only half the amount of histidine. Taurine and glycine, on the other hand, showed no significant changes throughout starvation. Taurine became the most predominant in the FAA pool after 40 days of starvation, and the concentration of 60-day-starved fish was two times higher than that of control group fish without starvation. The ratios of histidine, taurine, and glycine to total FAAs remained approximately the same although the individual contributions varied considerably to the total FAAs during starvation. The results of this study suggested that a good strategy would be to keep taurine and glycine in milkfish muscle at relatively high levels for physiological function as histidine decreased drastically for energy source under conditions of food deprivation.     

Histidine protects against zinc and nickel toxicity in Caenorhabditis elegans

Zinc is an essential trace element involved in a wide range of biological processes and human diseases. Zinc excess is deleterious, and animals require mechanisms to protect against zinc toxicity. To identify genes that modulate zinc tolerance, we performed a forward genetic screen for Caenorhabditis elegans mutants that were resistant to zinc toxicity. Here we demonstrate that mutations of the C. elegans histidine ammonia lyase (haly-1) gene promote zinc tolerance. C. elegans haly-1 encodes a protein that is homologous to vertebrate HAL, an enzyme that converts histidine to urocanic acid. haly-1 mutant animals displayed elevated levels of histidine, indicating that C. elegans HALY-1 protein is an enzyme involved in histidine catabolism. These results suggest the model that elevated histidine chelates zinc and thereby reduces zinc toxicity. Supporting this hypothesis, we demonstrated that dietary histidine promotes zinc tolerance. Nickel is another metal that binds histidine with high affinity. We demonstrated that haly-1 mutant animals are resistant to nickel toxicity and dietary histidine promotes nickel tolerance in wild-type animals. These studies identify a novel role for haly-1 and histidine in zinc metabolism and may be relevant for other animals.     

Enzymatic Production of Urocanic Acid by Achromobacter liquidum

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.     

Histidine utilization by Alcaligenes eutrophus: Regulation of histidase formation under heterotrophic conditions of growth

Histidine supported good growth of Alcaligenes eutrophus strain H 16 as a nitrogen source, but only poor growth as a carbon and energy source. The facultative chemolithoautotrophic bacterium was also able to utilize urocanic acid, the first intermediate of histidine catabolism. The products of histidine degradation were ammonium, formate and glutamate. Three enzymes of the pathway, histidase, urocanase and formiminoglutamate hydrolase, were present in histidine-grown cells. Two types of spontaneous mutants, derived from the wild type, were characterized by an increased growth rate on histidine. One of these types was found to produce histidase constitutively and at a higher activity compared with the parental strain. The second type of mutant had apparently gained an improved histidine uptake system, which is supposed to be growth rate-limiting in the wild type. From the physiological studies the conclusion was drawn that the control of histidine-degrading enzymes is based on induction by urocanate and catabolite repression by carbon sources supporting fast growth, such as succinate or pyruvate. Ammonium was found not to affect catabolite repression, however, we obtained evidence that histidine uptake is subject to a nitrogen control.Abbreviation CTAB hexadecyltrimethylammonium bromide     

2-fluorourocanic acid, a potent reversible inhibitor of urocanase.

The K_i for the interaction of 2-fluorourocanic acid with urocanase (from $\text{Pseudomonas fluorescens}$) is 1000 times as great as K_m for the natural substrate, urocanic acid, whereas enzymatic hydration of the fluoro analog occurs $\text{ca}$. 100 times more slowly. Inhibition is competive and is eventually overcome by utilization of the analog. By contrast, 4-fluoro- and 2-amino-urocanic acid are neither significant inhibitors nor substrates for the enzyme. 2-Fluorourocanic acid may prove a useful tool for blocking the utilization of histidine as a one-carbon source in metabolism.     

Urocanate reductase: identification of a novel anaerobic respiratory pathway in Shewanella oneidensis MR‐1

Interpretation of the constantly expanding body of genomic information requires that the function of each gene be established. Here we report the genomic analysis and structural modelling of a previously uncharacterized redox‐metabolism protein UrdA (SO_4620) of Shewanella oneidensis MR‐1, which led to a discovery of the novel enzymatic activity, urocanate reductase. Further cloning and expression of urdA, as well as purification and biochemical study of the gene's product UrdA and redox titration of its prosthetic groups confirmed that the latter is indeed a flavin‐containing enzyme catalysing the unidirectional reaction of two‐electron reduction of urocanic acid to deamino‐histidine, an activity not reported earlier. UrdA exhibits both high substrate affinity and high turnover rate (K_m << 10 μM, k_cat = 360 s^?1) and strong specificity in favour of urocanic acid. UrdA homologues are present in various bacterial genera, such as Shewanella, Fusobacterium and Clostridium, the latter including the human pathogen Clostridium tetani. The UrdA activity in S. oneidensis is induced by its substrate under anaerobic conditions and it enables anaerobic growth with urocanic acid as a sole terminal electron acceptor. The latter capability can provide the cells of UrdA‐containing bacteria with a niche where no other bacteria can compete and survive.     

In Vitro Catabolism of Histidine by Mixed Rumen Bacteriaand Protozoa

An in vitro study was conducted to examine the metabolism of histidine (His) by mixed rumen bacteria (B), mixed rumen protozoa (P), and a combination of the two (BP). Rumen microorganisms were collected from fistulated goats fed with lucerne cubes (Medicago sativa) and a concentrate mixture twice a day. Microbial suspensions were anaerobically incubated with or without 2 mm each of His, or histamine (HTM), or 1 mm urocanic acid (URA) at 39°C for 12 h. His and other related compounds in both supernatant and microbial hydrolysates were analyzed by HPLC. After 6- and 12-h incubations, the net degradation of His was 26.1% and 51.7% in B, 13.5% and 20.9% in P, and 21.7% and 46.0% in BP, respectively. The rate of the net degradation of His in B (98.0 μmol/g microbial nitrogen/h) was about 2.6 times higher than that of P during a 12-h incubation period. His was found to be degraded into urocanic acid (URA), imidazolelactic acid (ImLA), imidazoleacetic acid (ImAA), and histamine (HTM). Of these degraded His was mainly converted into URA in all microbial suspensions. The production of ImLA and ImAA was higher in B than in P suspensions, whereas the production of HTM was higher in P than in B suspensions. From these results, the existence of diverse catabolic routes of His in rumen microorganisms was indicated. Received: 23 May 2000 / Accepted: 31 July 2000     

Plasma amino acid levels in rainbow trout (Oncorhynchus mykiss)

The time course of plasma amino acid concentrations was studied in adult rainbow trout (300 g mean body weight). After a starvation period of 2 days fish were force-fed either with fish protein concentrate or a mixture of acidic casein and Na-caseinate at a rate of 0.32% CP (N' 6.25) of body weight. Peak levels occurred for feeding fish protein concentrate 6–12 h and for the casein mix 18 h post-feeding. The increase of the essential amino acids was closely correlated to the amino acid profile of the test proteins, whereas the concentration differences of the non-essential amino acids were at no time correlated to the amino acid pattern of fish protein concentrate or even negatively correlated in case of casein. The limiting amino acids in the test proteins were determined by ranking the average concentration increases (decreases) of the individual essential amino acids. Accordingly, arginine and histidine were most deficient in casein; in fish protein concentrate tryptophan seems to be the first limiting amino acid, followed by isoleucine.     

Simultaneous determination of stable isotopically labelled l-histidine and urocanic acid in human plasma by stable isotope dilution mass spectrometry

A capillary gas chromatographic—mass spectrometric method for the simultaneous determination of stable isotopically labelled l-histidine (l-[3,3-²H₂,1′,3′-¹⁵N₂]histidine, l-His-[M + 4]) and urocanic acid ([3-²H,1′,3′-¹⁵N₂]urocanic acid, UA-[M + 3]) in human plasma was developed using dl-[2,3,3,5′-²H₄,2′-¹³C,1′,3′-¹⁵N₂]histidine (dl-His-[M + 7]) and [2,3,5′-²H₃,2′-¹³C,1′,3′-¹⁵N₂]urocanic acid (UA-[M + 6]) as internal standards. l-Histidine and urocanic acid were derivatized to ^αN-(trifluoroacetyl)-^imN-(ethoxycarbonyl)-l-histidine n-butyl ester and ^imN-(ethoxycarbonyl)urocanic acid n-butyl ester. Quantification was carried out by selected ion monitoring of the molecular ions of the respective derivatives of l-His-[M + 4], dl-His-[M + 7], UA-[M + 3] and UA-[M + 6]. The sensitivity, specificity, precision and accuracy of the method were demonstrated to be satisfactory for measuring plasma concentrations of l-His-[M + 4] and UA-[M + 3] following administration of trace amounts of l-His-[M + 4] to humans.     

Control of histidase formation and improvement of urocanic acid production inSerratia marcescens

Summary In the wild-type strain ofSerratia mar-cescens Sr41, histidase was subjected to both cata-bolite repression (CR) by glucose and nitrogen re-pression (NtR) by excess NH⁺ ₄. CR-or NtR-re-sistant mutants for histidase were isolated from urocanase-deficient strain Ud-8, which was more sensitive to CR and NtR than wild-type strain. CR-resistant strain Uc1016 showed a high forma-tion of histidase as well as of D-serine deaminase in the presence of glucose. However, the addition of 2% (NH₄)₂SO₄ to the minimal medium de-creased the histidase formation to a large extent. NtR-resistant strain Gcl98 showed a high forma-tion of histidase as well as of adenylylated glu-tamine synthetase even in the presence of glucose and excess NH⁺ ₄. NtR-resistant strain Gc2031, isolated from urocanic acid-producing strain UdT122, produced urocanic acid more rapidly than strain UdT122 and in proportion to the rate of growth, although the maximum amounts of urocanic acid produced were the same. During cultivation, histidine accumulation was less and histidase activity was four times higher in strain Gc2031 than in the strain UdT122.     

Transcriptome atlas of eight liver cell types uncovers effects of histidine catabolites on rat liver regeneration

Eight liver cell types were isolated using the methods of Percoll density gradient centrifugation and immunomagnetic beads to explore effects of histidine catabolites on rat liver regeneration. Rat Genome 230 2.0 Array was used to detect the expression profiles of genes associated with metabolism of histidine and its catabolites for the above-mentioned eight liver cell types, and bioinformatic and systems biology approaches were employed to analyse the relationship between above genes and rat liver regeneration. The results showed that the urocanic acid (UA) was degraded from histidine in Kupffer cells, acts on Kupffer cells itself and dendritic cells to generate immune suppression by autocrine and paracrine modes. Hepatocytes, biliary epithelia cells, oval cells and dendritic cells can convert histidine to histamine, which can promote sinusoidal endothelial cells proliferation by GsM pathway, and promote the proliferation of hepatocytes and biliary epithelia cells by GqM pathway.