Search PubMed⌕ Search

Biomedical subjects

A Spisni

Publications and source records attributed to A Spisni.

At least 37 records · Page 2Linked to original sources
PubMed1989

Conformation of bombesin in buffer and in the presence of lysolecithin micelles: NMR, CD, and fluorescence studies.

The conformation of the tetradecapeptide hormone bombesin has been studied in buffer and in the presence of lysolecithin micelles, using static and dynamic fluorescence, CD, and one- and two-dimensional nmr. The results obtained show that in buffer bombesin is present in an extended flexible chain, with no evidence for any ordered secondary structure. A marked change in the CD spectrum is observed changing from buffer to the lipid suspension. Concomitantly, the 1H-nmr spectrum of bombesin, in a D2O lipid dispersion, shows the persistence of resonances due to exchangeable protons and in similar conditions the fluorescence intensity increases. We think therefore that these results strongly support the hypothesis that bombesin interacts with the lipid phase, assuming ordered secondary structure. Finally, the marked dependence of tryptophan fluorescence quantum efficiency and order parameter from the hormone concentration in the presence of lysolecithin but not in buffer leads to the conclusion that bombesin can associate into the lipid matrix.

Bombesin↗
PubMed1989

pH regulation of calcium recognition by an amino acid containing acyclic ionophore.

Calcium ion recognition by a dicarboxylic ionophore containing two (S)-phenylalanine residues joined via an amide bond by a flexible tri-oxa-undecanoyl bridge (Phe-3-O) has been investigated in a wide pH range (from pH 2 to 12). Experiments were performed in methanol and chloroform by 1H and 13C NMR, relaxation times and 2D NMR (COSY, NOESY and J-resolved experiments). Recognition is shown to be regulated by pH, as it occurs in at least three different coordination modes according to the experimental conditions. Even at low pH (pH 2) the ion is already complexed in the compartment created by the ethereal oxygens and the amide carbonyls. At higher pH, it becomes fully encapsulated in a pseudo-cyclic structure and at very basic pH it is localized between the amide carbonyls and the carboxylates. For these peculiar properties Phe-3-O appears to be a very promising ionophore in a trans-membrane pH gradient system.

Calcium↗
PubMed1988

Glycine transport by cultured human fibroblasts.

The transport of glycine was studied in cultured human fibroblasts. The amino acid entered the cell by Na+-dependent and Na+-independent mechanisms. Na+-independent glycine (0.1 mM) transport was less than 10% of total uptake and occurred by a mechanism formally indistinguishable from diffusion. Two distinct routes contributed to Na+-dependent glycine transport. The first route was identified with system A because it was inhibited by MeAIB and underwent adaptive regulation. The second route was identified with system ASC as it was inhibited by L-alanine, but not by MeAIB. Kinetic analysis revealed that the two systems operated glycine transport with the same Km of 1.6 mM, a value unusually high for system ASC.

Adult↗
PubMed1986

17O NMR investigation of the hydration of L-alanine and L-proline in water/Me2SO mixtures.

The hydration state of L-Alanine and L-Proline has been assessed via 17O NMR. At neutral and basic pH, two water molecules are hydrogen bonded at the carboxylate group, one to each oxygen, whereas a third water molecule is hydrogen bonded to the protonated COOH group at acidic pH, via the hydroxyl hydrogen. The possible formation of dimers and/or higher complexes in DMSO is indicated not only from the chemical shift but also from the linewidth of the amino acids.

Alanine↗
PubMed1985

Isolation and enzymatic characterization of the plasmalemma from bovine spermatozoa.

An improved method for the isolation of pure plasma and acrosomal membranes from bull spermatozoa is presented. Plasma membranes were isolated from the spermatozoa of bulls of different breeds, and some enzymatic activity, such as (Na+-K+) ATPase, Ca++ ATPase, Mg++ ATPase, alkaline and acidic phosphatases were assayed. Such enzymatic activity levels differ noticeably from those published by other authors, whose preparations were probably contaminated by other cellular components. Highly statistically significant differences of these activities have been found among the several breeds.

Acid Phosphatase↗
PubMed1984

Hypouricemic action of diflunisal in gouty patients: in vitro and in vivo studies.

Sixteen hyperuricemic gouty patients were treated with diflunisal, a novel salicylate, 500 mg BID. Serum and urine uric acid along with uric acid clearances were studied before and after a 7-day treatment. A clear hypouricemic action was observed, but 2 different mechanisms of action were seen when overexcretor patients and normoexcretor patients were analyzed separately. In overexcretors an allopurinol-like action was evident, whereas in normoexcretors a uricosuric action occurred. In vitro experiments showed a competitive inhibition of xanthine-oxidase exerted by diflunisal at low concentrations.

Adult↗
PubMed1983

Supramolecular organization of lysophosphatidylcholine-packaged Gramicidin A.

Heat derived gramicidin A'/L-alpha-lysophosphatidylcholine complexes were separated on a sucrose gradient to form two fractions: Fraction A which had an approximately constant Gramicidin A' to phospholipid ratio of 8 to 10 lipid molecules per Gramicidin A' molecule and Fraction B which had a larger but variable ratio. Fluorescence and circular dichroism studies confirmed Fraction A to be a lipid-incorporated channel state. Electron microscopic studies, using uranyl acetate negative staining, showed fraction A to be a membranous state with the formation of bilayer vesicles, that is, the interaction of peptide and phospholipid micelles causes the lipid to reorganize into a bilayer structure. Freeze-fracture replicas of the channel incorporated state demonstrated the presence of a supramolecular organization of particles exhibiting a tendency to form rows with a 50-60 A periodicity along the row and with 70-80 A distance between rows. An idealized working model for the incorporated state is presented.

Biological Transport↗
PubMed1983

Gramicidin A induces lysolecithin to form bilayers.

Heat-induced association of Gramicidin A with lysolecithin micelles results in the formation of lipid bilayer structures. The capacity of the Gramicidin A peptide to transform the lysolecithin lipid structure from micelle to bilayer is considered in terms of molecular packing mechanisms and relevance to membrane processes in general. The resulting lipid-bilayer-packaged channel system has particular usefulness in characterizing channel structure and mechanism.

Chemical Phenomena↗
PubMed1982

Antioxidant activity of bull semen in relation with aging.

This paper reports on the findings of an antioxidant activity in whole semen from bull and its components, washed spermatozoa and seminal fluid. The antioxidant activity has been evaluated as the ability of semen or its components to inhibit the spontaneous autooxidation of epinephrine a pH 10, 2, which involves the production of superoxide radicals (0(2)). This preliminary study provides further evidence on the role of free radicals, which are well known to be dangerous to cellular life, and points to the need of a better understanding of the role of antioxidant activities as a cellular protective mechanism.

Aging↗
PubMed1982

Intermolecular interactions of gramicidin A' transmembrane channels incorporated into lysophosphatidylcholine lipid systems.

Fluorescence studies are reported on gramicidin A' incorporated into lysophosphatidylcholine phospholipid structures. The shift in the emission maximum during incorporation and the quenching of fluorescence by I- and by acrylamide of the incorporated state obtained after prolonged heating are consistent with the presence of the channel state comprised of two single-stranded beta 6 -helices associated head-to-head (formyl end-to-formyl end). The quantum yield for the incorporated state, when gramicidin A' is within the lipid matrix, is very low and indicates the occurrence of intermolecular Trp-Trp interactions. Possible interactions between channels within the lipid matrix are discussed utilizing Trp-Trp contacts.

Gramicidin↗
PubMed1982

Serum antioxidant activity and related variables in rheumatoid arthritis. Behaviour during sulphydrylant treatment.

Some parameters (serum antioxidant activity, -SH groups, ceruloplasmin, copper) involved in the mechanisms of defence against oxygen free radicals were investigated in 25 rheumatoid patients and in 15 control subjects, matched for sex and age. When comparing the results obtained in the two groups, there is in rheumatoid arthritis a marked increase in the serum-specific antioxidant activity, related to ceruloplasmin, ESR, haemoglobin and Ritchie's index and a net decrease in the -SH groups inversely related to Ritchie's index and to ceruloplasmin. On analysing the sensitivity of the various parameters in discriminating between rheumatoid and healthy subjects, it is found that the -SH groups are the most sensitive, followed by the specific antioxidant activity, copper content, and ceruloplasmin. The evaluation of the same parameters in a group of 14 rheumatoid patients before and after 6 months of therapy with an antioxidant drug, viz. Tiopronin, confirms their usefulness in providing information regarding response to the treatment and on the recovery from the disease.

Adult↗
PubMed1981

Effect of CoQ homologues on the fluidity of phospholipid bilayers a studied by fluorescence polarization of perylene.

The effect of Coenzyme Q analogues on the fluidity of egg lecithin bilayers was studied by fluorescence polarization of perylene. The parameter was clearly affected by the side-chain length of the quinones, by their concentration, and by their redox state. With increasing concentration, short-chain quinones decreased the bilayer fluidity, and did so to an even greater extent in their reduced state. On the contrary, long-chain analogues fluidized the bilayer, the effect being independent of their redox state. Such observations might explain the lower efficiency of short-chain quinones, as compared to the long-chain analogues, in restoring, in vitro, the respiratory activity of CoQ2-depleted mitochondria.

Benz(a)Anthracenes↗
PubMed1980

The malonyl gramicidin channel: NMR-derived rate constants and comparison of calculated and experimental single-channel currents.

Malonyl gramicidin is incorporated into lysolecithin micelles in a manner which satisfies a number of previously demonstrated criteria for the formation of the transmembrane channel structure. By means of sodium-23 nuclear magnetic resonance, two binding sites are observed: a tight site and a weak site with binding constants of approximately 100 M-1 and 1 M-1 respectively. In addition, off-rate constants from the two sites were estimated from NMR analyses to be kofft congruent to 3 X 10(5)/sec and koffw congruent to 2 X 10(7)/sec giving, with the binding constants, the on-rate constants, kont congruent to 3 X 10(7)/Msec and konw congruent to 2 X 10(7)/Msec. Five different multiple occupancy models with NMR-restricted energy profiles were considered for the purpose of calculating single-channel currents as a function of voltage and concentration utilizing the four NMR-derived rate constants (and an NMR-limit placed on a fifth rate constant for intrachannel ion translocation) in combination with Eyring rate theory for the introduction of voltage dependence. Using the X-ray diffraction results of Koeppe et al. (1979) for limiting the positions of the tight sites, the two-site model and a three-site model in which the weak sites occur after the tight site is filled were found to satisfactorily calculate the experimental currents (also reported here) and to fit the experimental currents extraordinarily well when the experimentally derived values were allowed to vary to a least squares best fit. Surprisingly the "best fit" values differed by only about a factor of two from the NMR-derived values, a variation that is well within the estimated experimental error of the rate constants. These results demonstrate the utility of ion nuclear magnetic resonance to determine rate constants relevant to transport through the gramicidin channel and of the Eyring rate theory to introduce voltage dependence.

Colloids↗
PubMed1980

Conformational studies on the gramicidin A transmembrane channel in lipid micelles and liposomes.

The interaction of gramicidin A with lysolecithin micelles and with lecithin liposomes is demonstrated by circular dichroism to result in several metastable conformational states. A stable state can be obtained after extensive heating when the gramicidin A was added dry or in ethanol solution to the phospholipid dispersion but the stable state is readily obtained when gramicidin A is added in a trifluoroethanol solution. The circular dichroism of the stable conformational states is characterized by negative ellipticity below 205 nm and principally by a positive 220 nm band on which is superposed a weak 230 nm band (the latter likely arising from tryptophan side chains). The stable conformational state is considered to be that of the functional transmembrane channel primarily on the basis of extensive studies on its interaction with sodium ions.

Circular Dichroism↗
PubMed1980

Rate theory calculation of gramicidin single-channel currents using NMR-derived rate constants.

By means of 23Na NMR, two ion binding sites were observed in phospholipid-packaged gramicidin channels and the four associated rate constants were approximated. Limits also were placed on a fifth rate constant for an intrachannel ion translocation. By using Eyring rate theory to introduce voltage dependence, these rate constants were used in steady-state-current equations for calculation of gramicidin single-channel currents for two- and three-site models. Calculated single-channel currents are compared with previously published experimental single-channel currents obtained by electrical measurements on Na+ transport across gramicidin-doped planar lipid bilayers. The calculated results for the two- and three-site models compare favorably with the experimental results. Accordingly, it is demonstrated that NMR-derived rate constants can be coupled with Eyring rate theory to calculate currents through a transmembrane channel and to do so within levels of variation that compare with the differences obtained on planar lipid bilayers formed with different lipids.

Binding Sites↗
PubMed1979

Effect of phospholipase A2 on purified gastric vesicles.

The phospholipid and fatty acid composition and role of phospholipids in enzyme and transport function of gastric (H+ + K+)-ATPase vesicles was studied using phospholipase A2 (bee venom). The composition (%) was phosphatidyl-choline (PC) 33%; sphingomyelin (sph) 25%; phosphatidylethanolamine (PE) 22%; phosphatidylserine (PS) 11%; and phosphatidylinositol (PI) 8%. The fatty acid composition showed a high degree of unsaturation. In both fresh and lyophilized preparations, even with prolonged incubation, only 50% of phospholipids were hydrolyzed, but the amount of PE and PS disappearing was increased following lyophilization. There was a marked decrease in K+-ATPase activity (75%) but essentially no loss of the associated K+ p-nitrophenyl phosphatase was found. ATPase activity could be largely restored by various phospholipids (PE greater than PC greater than PS). There was also an increase in Mg2+-ATPase activity, partially reversed in fresh preparations by the addition of phospholipids (PE greater than PS greater than PC). Proton transport activity of the preparation was rapidly inhibited, initially due to a large increase in the HCl permeability of the preparation. Associated with these enzymatic and functional changes, the ATP-induced conformational changes, as indicated by circular dichroism spectra were inhibited.

Adenosine Triphosphatases↗