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Biomedical subjects

M Tabak

Publications and source records attributed to M Tabak.

At least 73 records · Page 4Linked to original sources
PubMed1992

Interaction of dipyridamole with micelles of lysophosphatidylcholine and with bovine serum albumin: fluorescence studies.

The interaction of the coronary vasodilator dipyridamole with biological systems, protein and membranes has been studied through optical absorption and fluorescence spectroscopies. Using the analysis of the spectra and fluorescence intensity of dipyridamole (DIP) in solution, the interaction of this compound with the transport protein albumin (BSA) and with a model of cell membranes, namely micelles of lysophosphatidylcholine (L-PC), was investigated. Measurements were performed at pH 5.0 and pH 7.0 where the molecule of DIP is fully protonated and partially protonated, respectively. The quenching of fluorescence with nitroxide-stable radicals 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) and 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPOL) as well as with acrylamide and iodide allowed the localization of the drug in the polar interface of micelles. Quenching by acrylamide and iodide in L-PC micelles demonstrated the effect of micelle protonation which increased the accessibility of iodide to the chromophore. An effective association constant was obtained both at pH 7.0 (7.5 x 10(3) M-1) and pH 5.0 (2.5 x 10(3) M-1) and a very good agreement with the proposed binding model was observed. The quantum yields of fluorescence data agree very well with the fluorescence lifetimes. The measurement of lifetimes was important to understand the kinetic data obtained from Stern-Volmer plots both of radical, acrylamide and iodide quenching of fluorescence. It was observed that, in the presence of micelles, the kq value increased for TEMPO while decreased for TEMPOL. This result, together with the vanishing solubility of DIP in saturated hydrocarbons and the preferential partition of TEMPO in micelles, suggested the localization of DIP in the polar micellar interface. This is also supported by the enhanced iodide quenching at pH 5.0, constancy of acrylamide quenching in the range of pH 7.0-5.0 and the partition of TEMPO and TEMPOL in SDS micelles. The association constant of DIP to BSA was also estimated both at pH 7.0 (2 x 10(4) M-1) and pH 5.0 (4 x 10(3) M-1). Quenching studies with nitroxide radicals, acrylamide and iodide also suggested the binding of the drug to a hydrophobic region of the protein. At pH 5.0, the protein undergo a conformational change which leads to a loosening of the overall structure so that the accessibility of the nitroxide radicals for DIP is increased at this pH. The differences in kq values at pH 7.0 and pH 5.0 suggested that at pH 7.0 the chromophore is protected in the protein site.(ABSTRACT TRUNCATED AT 400 WORDS)

Dipyridamole↗
PubMed1991

The lymphocyte transformation test with type II collagen as a diagnostic tool of autoimmune sensorineural hearing loss.

Immunological disorders of the cellular type can be diagnosed by the lymphocyte transformation test (LTT). An autoimmune mechanism of certain cases of sensorineural hearing loss (SNHL) can be evaluated by using human inner ear tissue as an antigen. Recent studies have shown that type II collagen plays an important role, not only in some autoimmune mediated rheumatoid diseases, but also as an antigenic substrate of inner ear tissue in autoimmune sensorineural hearing loss. This paper deals with results of the lymphocyte transformation test using type II collagen as antigen in 68 patients with progressive sensorineural hearing loss (PSNHL) and 68 healthy volunteers. Thirty-four patients showed a strong stimulation in the lymphocyte transformation test, in contrast to only four volunteers in the control group, two of whom had a history of rheumatoid arthritis.

Adult↗
PubMed1990

An ESR study of pathologic red blood cell membranes (RBCM).

A comparative study of red blood cell membranes from normal subjects and beta-thalassemia and sickle cell anemia patients was performed by spin labeling at the lipidic and protein phase. The results show that the quantity of bound spin label is the same for sickle, thalassemic, and normal membranes. The data from 5-doxyl stearic acid suggest an increase in fluidity for the thalassemic membrane.

Adult↗
PubMed1990

Effect of low mole fraction of trehalose dicorynomycolate from Corynebacterium diphtheriae on water permeability and electrical capacitance of lipid bilayer membranes.

The effect of incorporation of different proportions of trehalose dicorynomycolates (TDC) into lecithin bilayer membranes was studied. It was found that TDC, induces a 14% decrease of water osmotic permeability (42.6 +/- 3.9 to 36.8 +/- 2.7 microns/s) at 1.6 mole%, suggesting that this substance leads to an increase of the degree of packing of the constituent lipid molecules. A condensing effect of TDC was also apparent from membrane electrical capacitance (Cm) measurement. By incorporating TDC into bilayer membranes, the value of Cm experienced a decrease of 29% at 1.6% mole fraction. This finding was taken to reflect an increase in membrane thickness, known in many examples, to be related to the condensing effect.

Cord Factors↗
PubMed1990

Effect of hydration in metHb: reversible changes monitored by ESR of iron.

The dehydration of human and bovine methemoglobins was monitored using ESR spectroscopy of the iron signal. The interconversion of the Fe(III) signal between the high spin form (at g approximately 6) in solution and low spin form (at g approximately 2) was quantitatively studied as a function of hydration. The dehydration process leads also to a loss of paramagnetism resulting in the appearance of about 40% Fe(II) below 0.40 grH2O/grHb. The remaining 60% of Fe(III) ESR signal is distributed as the residual high spin form (at g approximately 6, 5%) and low spin form (hemichromes H and P, 55%). The formation of hemichrome P was explained as resulting from the coordination of the cysteine residue at beta 93 with the iron atom which follows the rupture of the proximal histidine bond. Experiments with hemoglobins where the sulphur atom of cysteine beta 93 was blocked (N-ethylmaleimide) did not showed the hemichrome P, confirming the involvement of the sulphur atom. This implies that the dehydration process induces displacements and torsion of the F helix, drastically changing the iron coordination at proximal site. In agreement with this proposition the Fe(II) symmetry is pentacoordinated with the disrupted bond to the proximal histidine at fifth coordination. This is also supported by ESR experiments with nitrosyl complex at low hydrations. All conformational changes were reversibly modulated by hydration degree and partially by lyophilization rate. A one-cycle dehydration of bovine hemoglobin followed by solubilization shows 100% reversibility of hemichrome P. Increasing the number of cycles of dehydration-hydration reduces the reversibility degree. With three cycles a reversibility of 70%-75% is observed. The level of 0.40 grH2O/grHb was the critical hydration for the molecules to return to aquo met form and correspond also to a minimal water content necessary to cover all protein surface as obtained from other techniques.

Animals↗
PubMed1988

pH-dependent phase transition of chlorpromazine micellar solutions in the physiological range.

The effects of pH and drug concentration on aggregation properties of chlorpromazine-HCl (CPZ) are examined. The critical micelle concentration (cmc) changes from 0.2 mM at pH 7.3 to 2 mM at pH 5.6 as estimated from the stearic acid spin label solubility measurements. For concentrations above the cmc CPZ micelles undergo a concentration-, temperature- and pH-dependent transition leading to phase separation. This phase transition is followed by a sudden increase of light scattering. The phase diagram pH vs. concentration is obtained by observation of the cloud point for concentrations ranging from 0.01 to 10 mM. The intramicellar environment is probed at pH ranging from 5.5 to 8.0 using a stearic acid spin label. The intramicellar compactness increases smoothly with increasing pH suggesting the weakening of polar heads repulsion due to charge decrease. The reported results indicate that pH effects are relevant and should be properly taken into account in the performance and interpretation of experiments with CPZ.

Chlorpromazine↗
PubMed1988

The mechanism of reaction of nitrosyl with met- and oxymyoglobin: an ESR study.

In this ESR work we have studied the pentacoordinate symmetry in horse, whale and sperm-whale myoglobin (Mb) in different physical states such as solution and powder. Experiments were performed in which the following parameters were varied: the sample temperature, pH, reaction time with NO, and NO concentration. The results enabled us to explain the NO reaction mechanism in the oxy and met forms of myoglobin. The study of powder samples at different degrees of hydration allowed us to identify the diamagnetic intermediate species existent in the reaction of NO with met-Mb proposed in the literature. The results presented explain adequately the pH effect and temperature dependence observed in the ESR spectra obtained using the met-Mb sample solutions from Sigma Chemical Co., which consist of a mixture (13%) of Mb-O2.

Electron Spin Resonance Spectroscopy↗
PubMed1988

An ESR study of nitrosyl-Aplysia brasiliana myoglobin and nitrosyl annelidae Glossoscolex paulistus erythrocruorin.

The nitrosyl derivatives of Annelidae Glossoscolex paulistus hemoglobin (an earth worm erythrocruorin (Ec AGp)) and Aplysia brasiliana myoglobin (Mb Apb) are studied using ESR spectroscopy. These two proteins have a quite similar ESR spectra at 100 K, but a different temperature behaviour. The temperature dependence of the nitrosyl Mb Apb spectrum is in good agreement with the Boltzmann distribution. In the case of nitrosyl-Ec AGp, the results are explained by the existence of two types of spectrum in thermodynamic equilibrium, with delta H = 9.08 kJ/mol, delta S = 47.15 J/mol and T1/2 = 193 K. There is a great similarity of the nitrosyl-Ec AGp spectra with those reported for elephant myoglobin, suggesting the presence of the same heme environment with a glutamine residue in the distal site. The pH dependence of the spectrum of nitrosyl-Mb Apb shows that the affinity of nitrosyl binding is higher at high pH (7.3) than at low pH (4.6). The ESR parameters are the same for these two pH values.

Animals↗
PubMed1988

Characterization of protein spin labeling by maleimide: evidence for nitroxide reduction.

A quantitative determination of maleimide spin label (MAL) binding in oxi and met hemoglobin (Hb) and bovine serum albumin are investigated using double integration to the ESR signal. This determination permitted the observation that a considerable fraction of MAL is reduced, losing its paramagnetism. Experiments using the same spin label with myoglobin and Hb with blocked-SH groups, where reduction was not observed, indicate the involvement of SH groups in the process. The 4-hydroxy-2,2,6,6-tetramethylpiperidino-1-oxyl spin label (which is not able to bind in the SH group) is reduced too, but the dependence on the molar ratio is different in comparison with the MAL case. In both cases the reduction percentage depends on the molar ratio spin label to protein and to the protein concentration. In order to obtain the total SH groups labeled (two in the Hb case) it is necessary to use an excessive amount of label (around 18:1) in the 0.5 mM Hb concentration.

Cyclic N-Oxides↗
PubMed1985

The acid-alkaline transition of a sea turtle myoglobin: coexistence at high pH of high- and low-spin forms.

The microenvironment of the iron in a sea turtle Dermochelys coriacea myoglobin is studied using the spectroscopic techniques EPR and optical absorption. Optical absorption spectra in the visible region suggest a great homology between turtle Mb and other myoglobins, such as those from whale, human and elephant. The pK of the acid-alkaline transition is 8.4 slightly lower than the pK of whale and equal to that of elephant myoglobin. The EPR spectrum at pH 7.0 is characteristic of a high-spin configuration with axial symmetry (gx = gy = 5.95). At higher pH, this signal changes in a way different from that observed for whale myoglobin. We observe for turtle Mb both the formation of a low-spin configuration with rhombic symmetry (gx = 2.56, gy = 2.20, gz = 1.90) and of a high-spin species with rhombic distortion (gx = 6.79, gy = 5.18, gz = 2.12). This suggests a lowering of symmetry at the haem, so that now the x and y directions are no more equivalent. This can be explained by amino acid substitution at the distal positions of haem or to off-axial positioning of distal residues. The coexistence at high pH (pH 11.0) of these two spin forms could be explained by the existence of two protein conformations, in which the crystal field splitting factor, delta, and the electron exchange energy are of the same order, allowing the presence of different configurations simultaneously. The presence of different kinds of haem is ruled out by the experiments with nitrosyl turtle Mb and turtle Mb-F showing spectra very similar to those of whale myoglobin. The pk of the acid-alkaline transition, 8.5, obtained from EPR spectra, agrees very well with results from optical absorption.

Animals↗
PubMed1985

On the interaction of Cu2+ with the heavy dipeptide Gly-Trp.

Distinct species are observed upon complexing of glycil-triptophan with Cu2+. The spectroscopic characterization of these complexes formed in different pH was made using visible light absorption (350-1100 nm) and electron paramagnetic resonance at room and liquid-nitrogen temperatures, with the samples in aqueous solution at the ratio of 10L:1M. Three species were identified in the following pH ranges: 4.0-6.0, 6.5-11.0, and above 12.00. The spectroscopic data and pK values of the Gly-Trp deprotonatable groups (in the presence of the metal) suggest that the complexes are CuL2(pH approximately 5.0), CuL(H2O). The complex above pH = 12.00 showed the bulky effect of the tryptophan side chain on the stereochemistry of the complex. The square planar symmetry is destroyed and a distorted tetahedral symmetry is achieved: the hyperfine parameter Az is reduced towards the value that occurs in blue proteins and the lowering of axial symmetry can be viewed by an increase in [gx-gy]. The tridentate complex CuL(H2O) was crystallized and single crystal measurements gave the molecular gyromagnetic tensor, but spin-spin interaction between neighbor ions masked the copper hyperfine interaction.

Chemical Phenomena↗
PubMed1985

On the interaction of small molecules with hemoglobin: orientational effects of hydration layers in protein crystal.

The spin label TEMPO does not show a binding to hemoglobin molecule in solution. In the crystal however the spin label is bound and a considerable anisotropy in the ESR spectra is observed similar to that with covalent spin labeling. Since TEMPO is a small spherical molecule the anisotropy should be a consequence of the ordering in the crystal packing observed for the hydration layers. Besides the anisotropic sites (one per asymmetric unit) an isotropic signal is apparent. The population of these sites is sensitive to the temperature and above around 30 degrees C a transition is observed of the label from the anisotropic site to the isotropic one. This is consistent with a change in the hydration structure above this temperature so that the spin label is sensitive to the reorganization of water or crystallization. Results of simulations based on the relaxation theory in liquids are compared for different hemoglobin physical states: crystal, powder and solution. It is shown that the ESR parameters obtained in the crystal are very different from those used for spectral stimulation at low temperature.

Crystallization↗
PubMed1980

Oxygen affinity of hemoglobin in polycythemia of undetermined etiology.

High oxygen affinity hemoglobinopathy, as an eventual cause for polycythemia of undetermined etiology, was investigated in 40 unclassifiable polycythemia subjects. The determination of 2,3-diphosphoglycerate and P50 in the polycythemic and in the control groups has not demonstrated any sigificant difference between them. The routine search of this condition in similar patients does not seem to be warranted.

2,3-Diphosphoglycerate↗
PubMed1978

The lipid-protein interaction in the membrane-bound Na+, K+-ATPase: a spin label study.

The interrelated temperature induced structural changes in the protein and lipid fractions of the Na+, K+-ATPase preparations were studied. A spin-labelled analog of ATP (ATP) was used, where the paramagnetic fragment was attached to the 2' (3')-OH ribose group. It is shown that the rotational mobility of ATP changes with the temperature in a discontinuous way. This correlates with the behaviour of the enzymatic activity of Na+, K+-ATPase and with the state of lipids in the enzyme preparations, both being characterized by a break in the 20-23 degrees C temperature region. When the Mn2+ ions were substituted by Mn2+ a strong magnetic dipolar interaction between the spin label in ATP and the Mn2+ ion was observed, which proves that the complex E-ATP--Mn2+ is formed before the hydrolysis of ATP. The structural changes which occur near 20 degrees C were also observed in the neighbourhood of the SH-groups modified by spin label X, an analog of maleimide. The structural changes observed support the idea that the protein-lipid interactions in the Na+, K+-ATPase provide the relaxation of the system from the unstable state during hydrolysis of ATP and cation transport.

Adenosine Triphosphatases↗
PubMed1977

[Interaction of a membrane preparation of Na+, K+-ATPase with a spin-labeled analog of ATP].

Interaction of membrane Na+, K+-ATPase preparation from brain gray matter with spin-labelled ATP analogue, in which free iminoxyl radical is joined as a result of 2'(3')-OH ribose groups acylation, is studied. The rotatory mobility of spin-labelled ATP analogue in Na+,K+-ATPase preparation is found to change in non-linear manner during temperature variation (the break-point on the curve being at 20-23degrees C). It correlates with temperature dependence of Na+,K+-ATPase and temperature dependence of lipid viscosity in the membranes, determined by means of hydrophobic spin probes. Substitution of Mg2+ ions with paramagnetic Mn2+ ions resulted in an intense magnetic dipole-dipole interaction between a spin label and Mn2+ ion, which indicated the formation of triple complex enzyme--spin-labelled ATP--Mn2+.

Adenosine Triphosphatases↗