Taurine Binding Research Articles

Calyculin A modulates the kinetic constants for the Na +-coupled taurine transport in Ehrlich ascites tumour cells

The effect of the phosphatase inhibitor calyculin A (cal A) on the kinetic parameters of the Na +-coupled taurine uptake via the taurine transporter in the Ehrlich ascites tumour cells has been investigated. Preincubation with cal A (100 nM) reduces the initial taurine influx by about 20%, but has no effect on the diffusional component of the taurine influx or on the taurine release from cells suspended in isotonic or in hypotonic medium. Thus, cal A-sensitive phosphatases only affect taurine transport mediated by the Na +-dependent taurine transporter. Cal A increases the Michaelis–Menten constant for binding of taurine to the transporter from 31±6 to 45±4 μM and reduces the taurine transport capacity from 210±20 to 170±10 pmol×g dry wt −1×min −1. The Michaelis–Menten constant for binding of Na + to the taurine transporter is concomitantly increased from 96±11 to 129±8 mM and the Na +:taurine coupling ratio for activation of the transport cycle is reduced from 3.3±0.6 to 2.4±0.2. This suggests that cal A-sensitive phosphatases maintain a high affinity of the taurine transporter towards Na + and taurine as well as a high taurine transport capacity in unpertubated Ehrlich cells.

Inhibition of taurine and 5?AMP olfactory receptor sites of the spiny lobster Panulirus argus by odorant compounds and mixtures

1. The effects of the odorant compounds adenosine-5'-monophosphate (5'AMP), ammonium, betaine, L-cysteine, L-glutamate, DL-succinate, and taurine and of mixtures of these compounds on binding of taurine and 5'AMP to dendritic membrane from the olfactory organ of spiny lobsters (Panulirus argus) were quantified to evaluate the contribution of inhibition of odorant-receptor binding to the generation of physiological responses to mixtures. 2. Taurine binding sites belong to two affinity classes, while 5'AMP binding sites belong to a single affinity class. Binding of either taurine or 5'AMP was partially inhibited in an apparently noncompetitive, concentration dependent fashion by most odorant compounds, with 25-40% inhibition by 1 mM of odorant. Mixtures of two or more odorant compounds also inhibited binding of taurine or 5'AMP to its sites. However, the inhibition by mixtures was often significantly less than expected from the inhibition produced by a mixture's components assuming either a noncompetitive or competitive mechanism. 3. By including this binding inhibition between compounds into models for predicting physiological responses to mixtures from the responses to the components, the predictive power of the models is significantly improved. This result strongly suggests that binding inhibition can influence the physiological responsiveness of chemoreceptor cells to mixtures.

Isolation of endogenous modulators for the GABAA and taurine receptors

Several endogenous brain substances which inhibit [3H]muscimol binding were isolated, and one of them has been purified to apparent homogeneity. The purification involved the extraction of brain tissue with water, followed by several steps of gel filtration column chromatography and high performance liquid chromatography (HPLC). The muscimol binding inhibitor (MBI) thus obtained appeared to be homogeneous as judged from the elution profile of an HPLC column, in which a symmetrical peak was obtained when the eluate was monitored at either 220 or 280 nm. Furthermore, the MBI activity coincided with the absorption peak. The purified MBI is not gamma-amino butyric acid (GABA), beta-alanine or taurine since these amino acids are clearly separated from the MBI in the purification procedures. The MBI has no effect on benzodiazepine (BZ) binding or glutamate binding to their respective receptors. However, the MBI is a more potent inhibitor for [3H]taurine binding than that of [3H]muscimol binding. The MBI appears to be a small molecule (< 2000 Da) that is heat and acid/base stable. The chemical nature of the MBI is currently under investigation.

Biochemical characterization of independent olfactory receptor sites for 5′-AMP and taurine in the spiny lobster

To understand the initial events in chemosensory transduction (i.e. binding of odorants to olfactory receptor sites), we have utilized a radioligand-receptor binding assay on a tissue preparation that is enriched in dendritic membrane from olfactory receptor cells in the spiny lobster Panulirus argus. Radioligands used were tritiated adenosine 5'-monophosphate (AMP) and taurine, which are two of the most excitatory compounds for spiny lobsters. Our results indicate the existence of two independent types of binding sites--a taurine binding site and an AMP binding site. For both the taurine and AMP binding sites, odorant binding is rapid, reversible, and saturable. KD values for the taurine and AMP binding sites are 2.3 and 2.0 microM, respectively, and Bmax values are 159 and 3.2 fmol/micrograms protein, respectively. The fact that the specificity, affinity, and independence of these two binding sites as defined in these biochemical studies are in agreement with those from electrophysiological studies suggests that these binding sites are olfactory receptor sites involved in sensory transduction.

Identification of the taurine binding proteins from human placental brush border membrane

Identification of the taurine binding proteins from human placental brush border membrane

Sodium-dependent and sodium-independent binding of taurine to rat brain synaptosomes

Rat brain synaptosomes (P 2B fraction) release μM concentrations of taurine into the incubation medium when incubated at protein concentrations of 0.5–5.0 mg/ml. Repeated incubations, with or without sonication, also induce release of μM concentration of taurine. When this release is taken into account, two classes of binding sites for taurine can be demonstrated over the intracellular-extracellular concentration range (3 μM–30 mM) in the presence of 140 mM sodium. The higher affinity class has a K d of 25.1 ± 7.7 μM (mean ± SE) and a B max of 15.08 ± 4.27 pmol mg −1 protein. This site is sodium-dependent, no binding being observed in the absence of sodium. Binding is also suppressed by the taurine transport inhibitor, guanidinoethane sulfonate. We equate this site with the transport site for taurine. The lower affinity site has a K d of 33.4 ± 7.5 mM and a B max of 10.3 ± 1.9 nmol mg −1 protein. Binding to this site is unaffected by guanidinoethane sulfonate. In the absence of sodium, K d and B max are decreased to 5.8 ± 0.56 mM and 2.55 ± 0.21 nmol mg −1 protein, respectively. We equate this site with the calcium-modulatory binding of taurine to phospholipid recently demonstrated in cardiac sarcolemma [Huxtable and Sebring, TIPS 7, 481 (1986)].

Taurine binding to the purified insulin receptor

Taurine (2-aminoethanesulfonic acid) was shown to bind specifically and reversibly to the purified human insulin receptor. While insulin binding to the purified insulin receptor exhibited characteristic negative cooperativity and an apparent dissociation constant ( K d of 1.2 × 10 −9 M, taurine binding was shown to exhibit positive cooperativity and had a lower affinity for the insulin receptor. The apparent K d for taurine binding to the purified insulin receptor was calculated to be 130 × 10 −9 M and the maximum number of binding sites ( B max ) was 1.6 nmol/mg receptor protein. Chromatographic data demonstrated that taurine binds to the 138,000 molecular weight subunit of the insulin receptor. Taurine binding to the receptor protein was displaced by either taurine or insulin. Anti-human insulin receptor sera prevented insulin or taurine from binding to the receptor. Taurine binding to the protein was pH dependent, and sulfur-containing taurine analogues were able to displace taurine from the insulin receptor. These data supported our previous in vivo and in vitro observations that the hypoglycemic properties of taurine appear to be mediated through an interaction of taurine with the insulin receptor.

Co-operativity in sodium-independent taurine binding to brain membranes in the mouse

Sodium-independent taurine binding to mouse brain membranes treated twice with Triton X-100 exhibited properties of positive co-operativity, suggesting that two or more taurine molecules interact at the binding site. The proposed taurine antagonist 6-aminomethyl-3-methyl-4H-1,2,4-benzothiadiazine-1,1 -dioxide and the new anticonvulsant taurine derivative taltrimide as well as glycine and GABA with their antagonists displaced taurine binding, strychnine being the most effective.

Effects of the anticonvulsant taurine derivative, taltrimide, on membrane transport and binding of GABA and taurine in the mouse cerebrum

The effects of a new anticonvulsive derivative of taurine, taltrimide (2-phthalimido-ethanesulphon-N-isopropylamide), and its metabolites on the release, uptake and binding of γ-aminobutyric acid (GABA) and taurine, were studied in the cerebrum of the mouse. The potassium-stimulated release of taurine from slices of cerebral cortex was inhibited in vitro by taltrimide and its dealkylated metabolite, whereas the stimulated release of GABA was significantly enhanced by both drugs. The uptake of taurine and GABA were not markedly affected. Both taltrimide and its dealkylated metabolite strongly inhibited the sodium-independent binding of taurine to synaptic membranes of brain, the effects on the binding of GABA being less pronounced. The actions on synaptic binding of taurine and on depolarization-stimulated release of GABA may be of significance for their anticonvulsant properties.

Taurine deficiency in the rat and cat: effects on neurotoxic and biochemical actions of kainate.

Taurine occurs at high concentration in the brain and has been suspected to be a major inhibitory transmitter. However, only a compound that fulfils all the criteria for transmitter identification can be considered to play this role. One important criterion is that the pharmacological actions of the proposed transmitter be mimicked by the endogenous substance. This, in fact, has been demonstrated for taurine only in the cerebellar cortex (16). Thus, the present state of knowledge does not support a general transmitter role for taurine. Since taurine affects so many different processes in the brain, a “neuromodulator” function has been proposed. The fact that an exogenously added agent influences a process is not necessarily relevant for its normal function. A better understanding of the physiological role for a substance may come from studies where the endogenous level of that compound has been lowered, and physiological perturbations are observed. One of the chief reasons why so little is known about taurine function is that this experimental approach is difficult. There are presently two ways to deplete endogenous taurine selectively. One is to administer the taurine uptake blocker 2-guanidinoethane sulfonate (GES) to rodents (5), and the other is to feed cats a taurine-deficient diet (21,23). The first approach is inexpensive and comparatively rapid, but involves introduction of a neuroactive agent (15) which accumulates in the brain (5,11,12) and may have side-effects. The second approach leads to a more severe taurine depletion, but it is expensive and time-consuming. In both cases the treatment has to be carried out chronically. Adaptive changes may be induced and might indeed be anticipated if taurine has an important function. One such possibly homeostatic alteration is the claimed enhancement of Na+-independent binding of taurine after GES treatment (11).

Taurine and GABA binding in mouse brain: effects of freezing, washing and Triton X-100 treatment on membranes.

The sodium-independent binding of taurine and GABA were determined with mouse brain membranes subjected to various freezing-thawing cycles, washing procedures and Triton X-100 treatments. The treated membranes were characterized by electron microscopy and determination of their content of endogenous free amino acids, e.g., taurine, GABA, glutamate, aspartate and glycine. The washings and the detergent gradually disrupted all synaptic structures, yielding empty-looking sealed membrane pouches of various sizes. Only traces of taurine, GABA, aspartate and glutamate were present in the Triton X-100-treated membranes, whereas these membranes still contained some glycine. Concomitantly, the affinity and maximal capacity of GABA binding increased, indicating that the treatments progressively removed some endogenous GABA binding inhibitors. Saturable taurine binding became detectable only after the Triton X-100 treatment, endogenous taurine, removable from membranes only with difficulty, obviously hampering binding measurements.

Actions of taurine on the GABA-benzodiazepine receptor complex solubilized from rat brain

The actions of taurine on the solubilized GABA-benzodiazepine receptor complex were investigated, and the results compared to those obtained with detergent-treated membrane-bound receptors. The receptor complex of adult rat brain was solubilized with Triton X-100 or CHAPS (3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulphonate). The properties of the solubilized GABA and flunitrazepam binding sites were similar to those in washed brain membranes. Taurine displaced GABA from its receptor sites and inhibited GABA stimulation of flunitrazepam binding to receptor complexes solubilized with Triton X-100. Thus the modulatory action of taurine on the receptor complex in washed membrane preparations was well preserved after this solubilization. No specific taurine binding to either Triton- or CHAPS-solubilized sample could be demonstrated.

Low affinity binding of taurine to phospholiposomes and cardiac sarcolemma

A sarcollema-enriched membrane fraction was prepared from the hearts of Sprague-Dawley rats and its ability to bind taurine (0.5–150 mM) was measured. In the absence of cations, the sarcolemma bound a maximum of 661 nmol taurine / mg protein, with a dissociation constant of 19.2 mM and a Hill coefficient of 1.9, indicating positive cooperativity. Scatchard analysis of taurine bindign to sarcolemma gave a bell-shaped curve. Neither β-alanine nor guanidinoethane sulfonate, inhibitors of taurine transport, affected the degree of taurine binding to sarcolemma. However, hypotaurine was an effective antagonist. Equimolar concentrations of Ca 2+, Na + or K + alro reduced taurine binding. Heterogeneous phospholipid vesicles of phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and phosphatidylserine (18:19:2:1) also bound taurine with positive cooperatively, yielding a bell-shaped Scatchard curve. The affinity of taurine for these mixed phospholipid vesicles was enhanced by the inclusion of cholesterol (50%). Taurine associated in a maximum ratio of 1:1 with homogeneous vesicles of phosphatidylcholine or phosphatidylserine. Vesicles of phosphatidylethanolamine bound taurine in a maximum of ratio of 2:1, whereas those of phosphatidylinositol bound significant amounts of taurine. These studies demonstrate a low affinity to sarcolemma of taurine at concentrations normally present in rat heart. Similar levels of binding were observed in phospholipid vesicles, suggesting that the interaction of taurine with biological membranes involves phospholipids.

Effects of intrahypothalamic kainic acid injection on taurine levels, binding and uptake

The neurochemical effects of unilateral intrahypothalamic injection of kainic acid on taurine levels and synaptosomal uptake and binding of taurine were investigated. Seven days after the kainic acid injections, there were no changes in either taurine uptake or binding. However, taurine levels were significantly increased by 54% over the control contralateral side. These data are consistent with the hypothesis that taurine is localized in glial cells: the increased levels being a result of gliosis after kainic acid injections.

Free amino acids and the uptake and binding of taurine in the central nervous system of rats treated with guanidinoethanesulphonate

The content of free amino acids in the central nervous system, the uptake of taurine by slices and synaptosomes and taurine binding to synaptic membranes have been studied on adult male Wistar rats after a prolonged administration of 2-guanidinoethanesulphonate with or without a taurine-free diet. The 2-guanidinoethanesulphonate administration both alone and together with a taurine-free diet considerably reduced taurine levels in all regions studied, viz. cerebrum, cerebellum, brain stem and spinal cord, but the contents of methionine and γ-aminobutyric acid were increased. On the normal diet the maximal taurine loss occurred after a 3-week 2-guanidinoethanesulphonate administration, when also the 2-guanidinoethanesulphonate accumulation in the cerebrum was largest. On the taurine-free diet the taurine levels were still low after the 4-week 2-guanidinoethanesulphonate administration when the taurine content in cerebral synaptosomes was also reduced to almost one half. 2-Guanidinoethanesulphonate treatment modified the uptake and binding processes ot taurine in the synaptosomal preparations. The maximal high-affinity taurine uptake was reduced, whereas the affnity of the specific sodium-independent binding sites for taurine increased. The results suggest that the neuromodulator properties of taurine could be affected by the reduced taurine levels and or by the accumulated 2-guanidinoethanesulphonate.

Synthesis and Anticonvulsant Properties of Some 2-Aminoethanesulfonie Acid (Taurine) Derivatives

□ A series of 2-acylaminoethanesulfonamides were synthesized by treating the corresponding sulfonyl chlorides with ammonia, a primary, or a secondary amine. A few compounds displayed marked anticonvulsant activity in mice when tested for their potency in the maximal electroshock seizure test. The piperidino, benzamido, phthalimido, and phenylsuccinylimido derivatives were active, whereas the succinylimido, saccharinylimido, and norbornendicarboxylimido compounds showed no activity. The interference with the sodium-independent taurine binding to mouse brain synaptic membranes was assessed to elucidate the possible mode of anticonvulsant action.

Taurine binding by rat retinal membranes

Retinal membrane preparations contain endogenous taurine which is difficult to remove. By repeatedly washing the membranes in buffer the taurine content was reduced from 11·7±2·5 to 2·5±0·5 nmol/mg protein. However, complete elimination of the endogenous taurine from the membrane preparation was not achieved. Binding of [ 3H]-taurine to rat retinal membrane preparations revealed both high and low affinity binding sites. A Hill coefficient of 0·71 suggested that cooperativity may be involved in the taurine binding process. Taurine binding was sodium dependent with maximum binding achieved at 118 m m. At suboptimal concentrations of sodium ions (30 m m) only one binding site was observed which appears to be the high affinity binding site. Analogues of taurine were tested for their effectiveness in displacing taurine from its binding site. Hypotaurine, β-alanine, γ-aminobutyric acid, and guanidinoethanesulfonic acid were the most potent displacers.

Sodium-independent taurine binding to brain synaptic membranes.

Saturable sodium-independent taurine binding to mouse and rat brain synaptic membranes was exposed after two freezing-thawing cycles combined with Triton X-100 treatments. The amount of saturable taurine binding was fairly low but was enhanced after depletion of brain taurine. Saturable taurine binding was displaceable by some convulsants and anticonvulsants but is specificity still remains to be established.

Neuronal [3H]benzodiazepine binding and levels of GABA, glutamate, and taurine are normal in Huntington's disease cerebellum.

Alterations in one subunit of the proposed GABA receptor complex, namely, the GABA receptor, have been observed in Huntington's disease cerebellum. We measured binding to a second subunit, the benzodiazepine binding site, in the autopsied cerebellum of 12 patients dying with adult-onset Huntington's disease. Neuronal benzodiazepine ([3H]flunitrazepam) binding density (Bmax) and affinity in cerebellar cortex of the Huntington's disease patients were not significantly different from control values. Similarly, maximal GABA stimulation of benzodiazepine binding was normal in the Huntington's disease cerebellum. In addition, no significant changes were observed in the concentrations of GABA, glutamate, and taurine in cerebellar cortex, nor of GABA in the dentate nucleus.

Anticonvulsant activity of some 2-aminoethanesulphonic acid (taurine) derivatives

A number of aliphatic 2-aminoethanesulphonic acid (taurine) derivatives were tested for their anticonvulsant activity in maximal electroshock seizure and pentylenetetrazole seizure threshold tests in mice. Certain piperidino, benzamido, phthalimido and phenylsuccinylimido derivatives were effective. Phthalimidoethanesulphon-N-methylamide, which inhibited the binding of both taurine and GABA to isolated synaptic membranes, appeared the best anticonvulsant among the derivatives tested.