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Biomedical subjects

I C Smith

Publications and source records attributed to I C Smith.

At least 163 records · Page 9Linked to original sources
PubMed1981

Effect of amphotericin B on membranes: a spin probe study.

The effect of the polyene antibiotic amphotericin B on the electron paramagnetic resonance spectra of lipid probes intercalated in model membranes was examined. When the antibiotic was added to the aqueous phase, no spectral effects occurred. However, when the antibiotic was incorporated during membrane preparation, changes in spectral parameters suggested the appearance of a new phase. The spectral changes do not necessarily corroborate the pore models proposed previously for amphotericin B in membranes. With a spin probe that partitions between water and membrane, an interaction between the amphotericin B and probe is observed. This interaction does not occur in the membrane, but in the aqueous phase, between the probe and the aggregated antibiotic. Some of the equilibria involving the antibiotic appear to be achieved slowly.

Amphotericin B↗
PubMed1981

Reliability of nitroxide spin probes in reporting membrane properties: a comparison of nitroxide- and deuterium-labeled steroids.

The reliability for the study of membrane properties of the steroid nitroxide spin probe, 3-doxylcholestane, was tested by comparison of analogous data for the deuterated steroid, cholesterol-3 alpha-d. Good agreement between the two probes was found for the dependence of their order parameters on variation of temperature or cholesterol concentration in egg phosphatidylcholine bilayers. This finding is contrasted with the results of a previous study of fatty acid probes where poor agreement was found for the spectral responses of nitroxide- and deuterium-labeled species. The angular dependence of the ESR spectra of nitroxide-labeled probes in oriented multibilayer films was examined to determine if the probes were oriented in a tilted fashion in the bilayer. The 3-doxylcholestane probe and a doxylstearic acid labeled at position 14 orient with their long molecular axes perpendicular to the bilayer plane. In contrast, the stearic acid probe nitroxide labeled at position 5 does not appear to orient in such a fashion. We suggest that the behavior of the latter probe reflects the difficulty of inserting a bulky nitroxide group into a highly ordered region of the bilayer rather than an inherent tilting of the phospholipid acyl chains. On the basis of the comparisons between various types of probes, some suggestions are made concerning the choice of ESR spin probe to obtain reliable information in membrane studies.

Cholesterol↗
PubMed1981

Kinetics of binding of methyl alpha- and beta-D-galactopyranoside to peanut agglutinin: a carbon-13 nuclear magnetic resonance study.

The binding kinetics of methyl alpha- and methyl beta-D-galactopyranoside to the anti-T lectin from peanuts were studied by 13C NMR, employing methyl galactopyranosides specifically enriched in 13C at C-1. Association and dissociation rate constants, as well as their activation parameters, are reported. The association rate constants, 4.6 X 10(4) M-1 s-1 for the alpha-galactopyranoside and 3.6 X 10(4) M-1 s-1 for the beta-galactopyranoside, are several orders of magnitude below those expected for a diffusion-controlled process. For both anomers, the association rate constant was temperature independent, implying that the association process occurs without a significant activation enthalpy. However, a considerable association activation entropy was found for both ligands. The dissociation rate constants were in the range of 9-46 s-1 within a temperature range of 5-35 degrees C for the alpha-galactopyranoside, and in the range of 9-39 s-1 within a temperature range of 5-25 degrees C for the beta-galactopyranoside. A considerable dissociation activation enthalpy of ca. 10 kcal mol-1 was found for both anomers. A two-step binding model, consistent with the present NMR data and with previous UV and CD spectroscopic data, is presented.

Circular Dichroism↗
PubMed1981

Novel polar lipids from the methanogen Methanospirillum hungatei GP1.

The methanogenic bacterium Methanospirillum hungatei GP1 has been shown to contain two unusual phosphoglycolipids (phosphoglycolipid I and phosphoglycolipid II) that account for 64% (by wt.) of the total cellular lipids. These lipids are derivatives of the dibiphytanyldiglycerol tetraether. One of the free hydroxyls of this tetraether is esterified with glycerophosphoric acid and the other is linked glycosidically to a disaccharide with structure alpha-Glcp-(1 leads to 2)-beta Gal phi in phosphoglycolipid I and beta-Gal phi-(1 leads to 6)-beta Gal phi in phosphoglycolipid II. Smaller amounts of the sn-2,3-diphytanylglycerol analog of phosphatidylglycerol and diglycosyldiphytanylglycerol ethers (DGD-I and DGD-II) containing the same disaccharide residues as in phosphoglycolipid I and phosphoglycolipid II, respectively, were identified, together with very small amounts of diglycosyldibiphytanyldiglycerol tetraethers (DGT-I and DGT-II) containing the same disaccharide residues as in phosphoglycolipid I and phosphoglycolipid II, respectively. A biosynthetic pathway involving head-to-head condensation of phosphatidylglycerol with DGD-I or DGD-II to form phosphoglycolipid I or phosphoglycolipid II, respectively, is proposed.

Carbohydrate Conformation↗
PubMed1981

Novel complex polar lipids from the methanogenic archaebacterium Methanospirillum hungatei.

The methanogenic archaebacterium Methanospirillum hungatei contains two unusual phosphoglycolipids that account for 64 percent of the total cellular lipids. These lipids are derivatives of the dibiphytanyl diglycerol tetraether, previously identified in methanogens. One of the free hydroxyls of this tetraether is esterified with glycerophosphoric acid, and the other is linked glycosidically to a disaccharide. The two phosphoglycolipids may function as covalently bonded lipid bilayers to impart stability and rigidity to methanogen membranes.

Cell Membrane↗
PubMed1981

Spin label reduction kinetics, a procedure to study the effect of drugs on membrane permeability: the effects of monosodium urate, dimethyl sulfoxide and amphotericin B.

A method is presented to study the effect of drugs on membrane permeability. It is based on the reduction of a spin label trapped in the internal aqueous compartment(s) of membranes by ascorbate ions added to the bulk aqueous phase. The decay of the electron spin resonance signal of the spin label as a function of time gives an indication of the effect of added agents on the permeability of membranes. To demonstrate the technique, the effect on model membranes of egg phosphatidylcholine of the gout-implicated compound monosodium urate, the aprotic solvent dimethyl sulfoxide and the polyene antibiotic amphotericin B were examined. Monosodium urate did not affect the permeability, casting doubt on a proposed mechanism whereby the agent disrupts the membranes via hydrogen bonding. Dimethyl sulfoxide promoted a gradual increase in rate of solute passage across cholesterol-containing model membranes. Amphotericin B had a pronounced effect on the permeability of cholesterol-containing membranes, causing nearly total loss of paramagnetism immediately after addition. Some aspects of the mechanism of action of the drugs are discussed as well as the advantages and disadvantages of the method. The experiments also allow the evaluation of the effect of surface charge and cholesterol on the dimensions of model membranes.

Amphotericin B↗
PubMed1981

Effects of amphotericin B on membrane permeability--kinetics of spin probe reduction.

The effect of the polyene antibiotic amphotericin B on the permeability of both unilamellar and multilamellar model membranes is investigated. The method measures the loss of the electron paramagnetic resonance signal of a spin probe, trapped in the aqueous compartment of a lipid dispersion, upon addition of ascorbate ions to the bulk aqueous phase. Amphotericin B causes large increases in the permeability of cholesterol-containing egg phosphatidylcholine membranes, whereas the effects are small in the absence of sterol and do not depend on surface charge. The effect of amphotericin depends upon the antibiotic:sterol mole ratio. The antibiotic appears to be unable to cross the membrane, acting only on the outermost bilayer of a multibilayer dispersion. When a phospholipid in the gel phase is used, amphotericin B causes large increases in permeability, independently of the presence or absence of sterol. It is suggested that the mechanism of action of amphotericin B is different for lipids in the liquid crystalline or gel states.

Amphotericin B↗
PubMed1981

The thermotropic behavior of dipalmitoyl phosphatidylcholine bilayers. A Fourier transform infrared study of specifically labeled lipids.

Fourier transform infrared spectroscopy has been used to study the thermotropic behavior of hydrated multibilayers of specifically deuterated derivatives of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine. It is shown that throughout the gel phase there is little or no conformational disorder introduced into the acyl chains. The pretransition effect is greatest in the central segments of the acyl chains, demonstrating that the interchain interactions are more pronounced in this region than in the center of the bilayer and suggesting the presence, in the gel phase, of a "plateau" in the strength of the interchain interactions. As the temperature is reduced, the rate of rotation of the terminal methyl group decreases steadily; below 0 degree C the conformation is rigid on the infrared time scale.

Lipid Bilayers↗
PubMed1981

Analysis of the composition of mixed lipid phases by the moments of 2H NMR spectra.

The use of (2)H NMR spectral moments to determine the composition of biphasic lipid mixtures is outlined. The analysis has been applied to phosphatidylethanolamine-cholesterol (1:1), potassium palmitate, 30% (wt/wt) water and phosphatidylcholine-cholesterol (4:1) systems, as well as to membrances of Escherichia coli during phase transitions. The advantages and disadvantages of the use of spectral moments to determine fractions of coexistent phases are discussed.

Cell Membrane↗
PubMed1980

Orientational order of unsaturated lipids in the membranes of Acholeplasma laidlawii as observed by 2H-NMR.

Oleic acid specifically deuterated at fifteen different positions along the chain, including the double bond, was biosynthetically incorporated into the membrane lipids of the microorganism Acholeplasma laidlawii B. A detailed study of the dynamic conformation of these chains was carried out using deuterium nuclear magnetic resonance. The deuterium spectra fourteen different samples were recorded as a function of temperature over the range 0-41 degrees C. Spectra were obtained down to -52 degrees C for the sample enriched with oleic acid deuterated at the C-12' position. Above 20 degrees C, where the lipids are in the liquid crystal phase, a single quadrupolar powder pattern was observed for each C2H2 segment, except for the C-2' position which gave rise to a three-component spectrum characteristic for this position in both model and biological membranes. Simulation of this spectrum indicates that there are two conformations of the lipid molecule in the region of the C-2' segment of the sn-2 chain. The orientationa fluctuations of the fatty acid chain segments in the A. laidlawii membranes are described by the deuterium order parameters, and a striking similarity is shown to exist between the oleate chain conformation of the A. laidlawii membrane and a phospholipid model membrane. Remarkable similarities are also demonstrated in the A. laidlawii membrane enriched in palmitic and oleic fatty acids when the order parameter profiles are plotted at the same reduced temperature. Below 15 degrees C a second component, due to gel phase lipid, starts to appear in the spectra. This broad gel phase component grows at the expense of the liquid crystal phase component as the temperature is reduced. The spectra indicate that the center of the phase transition is at about -12 degrees C, in good agreement with DSC studies.

Acholeplasma laidlawii↗
PubMed1980

The temperature dependence of molecular order and the influence of cholesterol in Acholeplasma laidlawii membranes.

2H nuclear magnetic resonance (NMR) of Acholesplasma laidlawii membranes grown on a medium supplemented with perdeuterated palmitic acid shows that at 42 degrees C or above, the membrane lipids are entirely in a fluid state, exhibiting the characteristic 'plateau' in the variation of deuterium quadrupolar splitting with chain position. Between 42 and 34 degrees C there is a well-defined gel-to-fluid phase transition encompassing the growth temperature of 37 degrees C, and at lower temperatures the membranes are in a highly ordered gel state. The 2H-NMR spectra of the gel phase membranes are similar to those of multilamellar dispersions of chain perdeuterated dipalmitoyl phosphatidylcholine (Davis, J.H. (1979) Biophys. J. 27, 339) as are the temperature dependences of the spectra and their moments. The incorporation of large amounts of cholesterol into the membrane removes the gel to fluid phase transition. Between 20 and 42 degrees C, the position dependence of the orientational order of the hydrocarbon chains of the membranes is similar to that of the fluid phase of the membranes without cholesterol, i.e., they exhibit the plateau in the deuterium quadrupolar splittings. However, the cholesterol-containing membranes have a higher average order, with the increases in order being greater for positions near the carbonyl group of the acyl chains. Below 20 degrees C the 2H spectra of the membranes containing cholesterol change dramatically in a fashion suggestive of complex motional and/or phase behaviour.

Acholeplasma laidlawii↗
PubMed1980

Acholeplasma laidlawii membranes: a Fourier transform infrared study of the influence of protein on lipid organization and dynamics.

Fourier transform infrared spectroscopy is applied to the study of intact and deproteinated plasma membranes of Acholeplasma laidlawii, enriched biosynthetically with perdeuteriopalmitoyl chains. The temperature-dependent behavior is monitored via the CD stretching modes and compared with that observed in the model membrane 1,2-diperdeuteriopalmitoyl-sn-glycero-3-phosphocholine. The broad ramplike transition observed in the natural membranes is shown to consist of two overlapping stages. In the lower temperature range the principal change is a reduction in the rigidity of the lipid matrix. Subsequently, over a 5 degrees C range, centered at the growth temperature, a large change in the gauche/trans conformer ratio of the acyl chains occurs, similar to that generally observed in model systems. This is followed by an abrupt cessation of the phase transition. The effects of membrane protein on the phase transition are shown to be relatively minor. Firstly, they produce a decrease in the rate of acyl chain motion at a given temperature, resulting in a reduction in the width of the transition. In addition, the presence of protein increases the population of gauche conformers of the fatty acyl chains in the liquid-crystalline phase of the membrane lipids.

Acholeplasma laidlawii↗
PubMed1980

Interaction of opioid peptides with model membranes. A carbon-13 nuclear magnetic study of enkephalin binding to phosphatidylserine.

The binding of enkephalin to phosphatidylserine has been studied, by using 13C NMR, as a model for interactions with components of biological receptors. Chemical shifts, line widths and spin-lattice relaxation times were measured for peptides enriched to 90% in 13C. The pKa values of the terminal amino and carboxyl groups were determined from the pH dependence of the 13C chemical shifts. Interaction of (2-[2-13C]glycine) methionine-enkephalin, (3-[2-13C]-glycine)methionine-enkaphalin, and (3-[2-13C]glycine)methionine-enkephalinamide with phosphatidylserine (PS) was studied as a function of pH. Salt and morphine antagonism to binding was manifest. Binding was shown to be pH dependent, exhibiting a maximum under slightly acidic conditions. Whereas the -NH3+ group of enkephalin is essential for binding, the data suggest that neither the tyrosyl hydroxyl group nor the COO- group is involved. Binding affects the 13C spin-lattice relaxation times most strongly; the chemical shifts and line widths of the 13C-enriched material show little perturbation in the presence of PS. The internal flexibility of the peptides is decreased, on binding to model membranes, by 1 order of magnitude. Dissociation constants have been measured as 4 X 10(-1) M and 2.6 X 10(-3) M for enkephalin and enkephalinamide, at pH 6.3 and 6.4, respectively.

Animals↗