Effects of phorbol ester and dexamethasone treatment on histidine decarboxylase and ornithine decarboxylase in basophilic cells

Both histamine and polyamines are important for maintaining basophilic cell function and viability. The synthesis of these biogenic amines is regulated by histidine decarboxylase and ornithine decarboxylase, respectively. In other mammalian tissues, an interplay between histamine and polyamine metabolisms has been suspected. In this report, the interplay between histamine and ornithine-derived polyamines was studied in a non-transformed mouse mast cell line (C57.1) treated with phorbol ester and dexamethasone, a treatment previously used to increase histidine decarboxylase expression in mastocytoma and basophilic leukemia. Treatment with phorbol ester and dexamethasone increased histidine decarboxylase expression and intracellular histamine levels in C57.1 mast cells to a greater extent than those found for other transformed basophilic models. The treatment also induced a reduction in ornithine decarboxylase expression, intracellular polyamine contents, and cell proliferation. These results indicate that the treatment induces a co-ordinate response of polyamine metabolism and proliferation in mast cells and other immune-related cells. The decrease in the proliferative capacity of mast cells caused by phorbol ester and dexamethasone was simultaneous to an increase in histamine production. Our results, together with those reported by other groups working with polyamine-treated mast cells, indicate an antagonism between histamine and polyamines in basophilic cells.     

Inhibition of human erythrocyte orotidylate decarboxylase

Ribonucleotide derivatives of allopurinol and oxipurinol are potent inhibitors of human erythrocyte orotidylate decarboxylase. The inhibition constants are dependent upon the aggregation state of the enzyme, much tighter binding being observed with higher molecular weight forms. The trend was similar to that observed for K_mvalues for orotidine-5'-phosphate, the substrate of the enzyme. Of the compounds tested, 1-oxipurinol-5'-phosphate, with a K_i, value of 0.3 nM for the 250,000 M.W. species, was the most effective inhibitor. This was some two orders of magnitude tighter than 7-oxipurinol-5'-phosphate, which in turn was two orders of magnitude tighter than 1-allopurinol-5'-phosphate. A similar trend of K_i estimates with molecular weight of the enzyme was observed with a number of other inhibitors, including 3-XMP, 9-XMP, 6-azaUMP, UMP and inorganic phosphate (HPO₄^2?).     

海藻酸钠法固定化谷氨酸脱羧酶的研究

目的研究海藻酸钠法固定化谷氨酸脱羧酶的最适条件。方法以海藻酸钠为载体,分别研究了海藻酸钠浓度、CaCl2浓度、固定化时间和固定化酶的最适pH、最适温度、最适底物浓度及反复利用的稳定性。结果最适固定化条件为3%海藻酸钠、0.1 mol/L CaCl2、固定化时间6 h。固定化酶的最适pH为5.0、最适温度为42℃、最适底物浓度1%,反应进行4次后一半的酶活损失,底物转化率在90%以上。结论海藻酸钠法固定化谷氨酸脱羧酶可以很好的转化谷氨酸钠生成γ-氨基丁酸,具有实用的潜力。     

Activity of aromatic-L-amino-acid decarboxylase in pheochromocytoma cells

The activity of aromatic-L-amino-acid decarboxylase in pheochromocytoma cells was assayed by measuring the formation of ¹⁴CO₂ by cells incubated with L-[1-¹⁴C]dopa. The V_max of dopa decarboxylation in intact cells was about 3 nmoles CO₂ produced/min per mg protein, and the apparent K_m of this process was approximately 32μM. The V_max of decarboxylase activity was at least an order of magnitude greater than the rate of dopa synthesis in these cells. Incubation of the cells with cholera toxin or with 56 mM K⁺ caused an activation of tyrosine 3-monooxygense and an increase in the rate of dopa production, but it did not affect the activity of the decarboxylase in the cells. The pheochromocytoma ocytoma cells contained about 12 pmoles dopa/mg protein. Treatment of the cells with cholera toxin or with high K⁺ increased the dopa content of the cells. Aromatic-L-amino-acid decarboxylase was not the rate-limiting enzyme in the pathway of catecholamine synthesis in these sells; moreover, it appeared that the activities of this enzyme and of tyrosine 3-monooxygenase were not regulated coordinately.     

Peptide inhibition of mammalian histidine decarboxylase.

The hypothesis that N-terminal histidine peptides might act as inhibitors to histidine decarboxylase was investigated. A murine mastocytoma was utilized as enzyme source. The crude extract of this tissue exhibits high rates of decarboxylation of both histidine and DOPA and was used to establish the specificity in the effect of the compounds tested. For kinetic analyses a highly purified histidine decarboxylase fraction was used. The effect of some representative peptides on both enzyme activities were recorded. Histidine decarboxylase exclusively was inhibited by N-terminal histidine peptides. None of the other peptides investigated interfered negatively with this enzyme. This inhibition was consistent in the purified preparation and appeared to be more pronounced with increasing hydrophobicity in the second amino acid. Histidyl-phenylalanine was found to be about 100-fold as potent as the commonly used specific histidine decarboxylase inhibitor alpha-methyl histidine. It is concluded that small peptides with histidine as the N-terminal amino acid might act as specific inhibitors for mammalian histidine decarboxylase. An analog effect of small tyrosyl or phenylalanyl peptides was not seen for the DOPA decarboxylase.     

Subcellular distribution of mouse mevalonate pyrophosphate decarboxylase

Mevalonate pyrophosphate decarboxylase (MPD) is considered to be a cytosolic protein. Recently, other groups reported that MPD is mostly located in the peroxisomes. In this study, we examined whether the expression of MPD in mice depends on the proliferation of peroxisomes, and whether MPD is predominantly located in the peroxisomes or the cytosol of mice. No increase in the protein level of MPD was observed in the crude extract of the livers of mice administered with peroxisome proliferative drugs. The result suggests that the expression of MPD is independent of the proliferation of peroxisomes, and may be maintained via a specific regulatory mechanism, different from the regulation of the expression of peroxisome proliferator-activated receptor alpha. When the subcellular distribution of MPD in mouse melanoma (B16F10) cells was examined by cell fractionation, MPD was detected in the cytosol of B16F10 cells, but not in the peroxisomes. In permeabilized B16F10 cells treated with digitonin, which lack cytosolic enzymes, 80% and 20% of MPD, 75% and 25% of lactate dehydrogenase, or 2% and 98% of catalase, existed in the medium and in the cell, respectively. From these results, it indicated that MPD was predominantly located in the cytosol and did not exist in the peroxisomes of B16F10 cells.     

Aromatic L-amino acid decarboxylase in calcitonin-producing cells

A common feature of the calcitonin-producing cells (C cells) is their capacity to produce and store arylethylamines. The activity of aromatic l-amino acid decarboxylase (DOPA/5-HTP decarboxylase) was measured radiometrically in the thyroid glands of various species and in the ultimobranchial gland of the chicken. The enzyme activity was well correlated with the number of amine-containing C cells, demonstrated by fluorescence microscopy in these tissues. The ultimobranchial gland had a conspicuously high activity of aromatic amino acid decarboxylase. The follicular cells of the thyroid appeared to have no or only a very low activity of this enzyme.     

A study of potential histidine decarboxylase inhibitors

A series of compounds has been examined for ability to inhibit histidine decarboxylase. Histidine analogues having substituents in the imidazole ring showed a wide variation in potency, but these were all much less active than α-methyldopa [β-(3,4-dihydroxyphenyl)-α-methylalanine], the most potent known inhibitor of histidine decarboxylase. Some tentative conclusions are drawn regarding the relationship between chemical structure and inhibitory activity.