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At least 1,171 records · Page 65Linked to original sources
PubMed1974

Translational diffusion coefficient and partition coefficient of a spin-labeled solute in lecithin bilayer membranes.

The translational diffusion coefficient and the partition coefficient of a spin-labeled solute, di-t-butyl nitroxide, in an aqueous suspension of dipalmitoyl lecithin vesicles have been studied by electron spin resonance spectroscopy. When the lecithin is cooled through its phase transition temperature near 41 degrees C, some solute is "frozen out" of the bilayer, and the standard partial molar enthalpy and entropy of partition go more positive by a factor of 8 and 6, respectively. However, the apparent diffusion constant in the lecithin phase is only slightly smaller than that in water, both above and below the transition temperature. The fraction of bilayer volume within which solute is distributed may increase with temperature, contributing to the positive enthalpy of partition. Comparison of time constants suggests that there is a permeability barrier to this solute in the periphery of the bilayer.

Butanes↗
PubMed2005

Effects of the apparent transverse relaxation time on cerebral blood flow measurements obtained by arterial spin labeling.

Previous modeling studies have predicted that a significant fraction of the signal in arterial spin labeling (ASL) experiments originates from labeled water in the capillaries. Provided that the relaxation times in blood and tissue are similar, ASL data can still be analyzed with the conventional one-compartment Kety model. Such studies have primarily focused on T1 differences and have neglected any differences in transverse relaxation times (T2 and T2*). This is reasonable for studies at lower fields; however, it may not be valid at higher fields due to the stronger susceptibility effects of deoxygenated blood. In this study a tracer kinetic model was developed that includes T2* differences between capillary blood and tissue. The model predicts that a reduction in blood T2* at higher fields will attenuate the capillary contribution to the ASL signal. This in turn causes an underestimation of CBF when ASL data are analyzed with the one-compartment Kety model. We confirmed this prediction by comparing ASL data collected at 1.5 and 4 T, and at multiple gradient echoes (19, 32, 45, and 58 ms). A decrease in resting-state CBF with echo time (TE) was observed at 4 T, but not at 1.5 T. These results suggest that at higher fields AST data should be collected using gradient-echo techniques with short TEs, or with spin-echo techniques. Furthermore, the sensitivity of the CBF measurements to venous T2* may affect the interpretation of concurrent ASL/BOLD studies.

Adult↗
PubMedDate not supplied

Nitroxyl spin label contrast enhancers for magnetic resonance imaging. Studies of acute toxicity and mutagenesis.

Two nitroxyl spin label (NSL) compounds that are used experimentally as in vivo contrast enhancers in magnetic resonance (MR) imaging were tested for acute toxicity in rats and for genotoxic effects in cell cultures. These compounds, 2,2,5,5-tetramethyl-1-pyrrolidinyl-oxy-3-carboxylic acid (PCA) and 2,2,6,6-tetramethyl-1-oxido-4-piperidinyl-1-succinic acid (TES) and their hydroxylamine and amine derivatives did not induce sister chromatid exchanges or mutations in Chinese hamster ovary cells at the HGPRT or Na+/K+ ATPase loci. The acute LD50 doses in rats for PCA and TES are 15.1 mmol/kg or greater, suggesting relatively high tolerance.

Animals↗
PubMed2004

Internal carotid artery occlusion assessed at pulsed arterial spin-labeling perfusion MR imaging at multiple delay times.

Magnetic resonance (MR) imaging with pulsed arterial spin labeling (ASL) was performed at six different inversion times in nine patients with internal carotid artery (ICA) occlusion and in 11 control subjects. The hospital's commission on scientific research on human subjects approved the study protocol, and all study subjects gave informed consent. Cerebral blood flow (CBF) in the middle cerebral artery territories was calculated from the combined signal intensities measured with ASL at the multiple inversion times. In the patients with ICA occlusion, mean CBF values were decreased in the gray matter of the hemisphere ipsilateral to the occlusion, as compared with values in the gray matter of the contralateral hemisphere (P < .05) and with values in the gray matter of the control subjects (P < .05). Quantification of CBF with ASL at multiple inversion times can compensate for the blood transit delays in patients with ICA occlusion.

Adult↗
PubMed2002

Rapid transbilayer movement of spin-labeled steroids in human erythrocytes and in liposomes.

The transbilayer movement and distribution of spin-labeled analogs of the steroids androstane (SLA) and cholestane (SLC) were investigated in the human erythrocyte and in liposomes. Membranes were labeled with SLA or SLC, and the analogs in the outer leaflet were selectively reduced at 4C using 6-O-phenylascorbic acid. As shown previously, 6-O-phenylascorbic acid reduces rapidly nitroxides exposed on the outer leaflet, but its permeation of membranes is comparatively slow and thus does not interfere with the assay. From the reduction kinetics, we infer that transbilayer movement of SLA in erythrocytes is rapid at 4C with a half-time of approximately 4.3 min and that the probe distributes almost symmetrically between both halves of the plasma membrane. We have no indication that a protein-mediated transport is involved in the rapid transbilayer movement of SLA because 1) pretreatment of erythrocytes with N-ethyl maleimide affected neither flip-flop nor transbilayer distribution of SLA and 2) flip-flop of SLA was also rapid in pure lipid membranes. The transbilayer dynamics of SLC in erythrocyte membranes could not be resolved by our assay. Thus, the rate of SLC flip-flop must be on the order of, or even faster than, that of probe reduction rate on the exoplasmic leaflet (half-time approximately 0.5 min). The results are discussed with regard to the transbilayer dynamics of cholesterol.

Androstanes↗
PubMed2003

Transient kinetics and mechanics of myosin's force-generating rotation in muscle: resolution of millisecond rotational transitions in the spin-labeled myosin light-chain domain.

We have used electron paramagnetic resonance (EPR) of spin-labeled scallop muscle, in conjunction with laser flash photolysis of caged ATP, to resolve millisecond rotational transitions of the myosin light-chain domain (LCD) during transient force generation. We previously used EPR to resolve two distinct orientations of the LCD [Baker, J. E., Brust-Mascher, I., Ramachandran, S., LaConte, L. E., and Thomas, D. D. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 2944-2949], correlated these structural states with biochemical states in the actin-myosin ATPase reaction, and showed that a small shift in the steady-state distribution between these two LCD orientations (i.e., a net lever arm rotation) is associated with force generation in muscle. In the study presented here, we measured millisecond changes in this orientational distribution (i.e., the rates of transition between the two LCD orientations) in muscle following flash photolysis of caged ATP, in both the presence and absence of Ca. The transient acquired in the absence of Ca is dominated by a rapid (1/tau(1) = 37 s(-1)) disordering transition from the single orientation in rigor to the bimodal orientation distribution observed for detached cross-bridges in relaxation (i.e., the reversal of the lever arm rotation), followed by a recovery phase (1/tau(2) = 2.4 s(-1)) of very small amplitude (small fraction of heads participating). In the presence of Ca, the transient exhibited a similar initial disordering phase (1/tau(1) = 38.5 s(-1)), followed by a recovery phase (1/tau(2) = 8.33 s(-1)) of substantial amplitude, corresponding to the forward rotation and ordering of the lever arm. A standard kinetic model was used to fit these data, revealing rate constants consistent with those previously determined by other methods. Surprisingly, a comparison of the EPR transients with force transients reveals that the rate of force development (91 s(-1)) is faster than the rate of the forward lever arm rotation (8 s(-1)). This observed relationship between the kinetics of the lever arm rotation and transient force development in muscle provides new insight into how myosin both generates and responds to muscle force.

Actins↗
PubMed1986

Investigation of surface potential asymmetry in phospholipid vesicles by a spin label relaxation method.

In earlier work, Castle and Hubbell (1976) demonstrated the use of a spin-labeled amphiphile as a probe for the electrostatic potential at the outer surface of charged phospholipid vesicles. In recent experiments, we have shown that the hydrophobic anion tetraphenylboron (TPB) promotes transbilayer migration of the probe molecule. Relaxation data recorded following the rapid mixing of the probe with TPB-containing vesicle samples provides information about the electrostatic potentials at both the outer and inner vesicle surfaces. The measured potentials for both surfaces of asymmetrically screened vesicles were found to be in good agreement with theoretical values calculated using their known surface charge density. The method is also sensitive to transmembrane potentials as indicated by the response of the label to potentials created with the use of potassium concentration gradients and valinomycin.

Cyclic N-Oxides↗
PubMed1988

A comparative spin-label study of isolated plasma membranes and plasma membranes of whole cells and protoplasts from cold-hardened and nonhardened winter rye.

Lipid-lipid and lipid-protein interactions in the plasma membranes of whole cells and protoplasts and an isolated plasma membrane fraction from winter rye (Secale cereale L. cv Puma) have been studied by spin labeling. Spectra were recorded between -40 degrees C and 40 degrees C using the freely diffusing spin-label, 16-doxyl stearic acid, as a midbilayer membrane probe. The probe was reduced by the whole cells and protoplasts and reoxidized by external potassium ferricyanide. The reoxidized probe was assumed to be localized in the plasma membrane. The spectra consisted of the superposition of a narrow and a broad component indicating that both fluid and immobilized lipids were present in the plasma membrane. The two components were separated by digital subtraction of the immobilized component. Temperature profiles of the membranes were developed using the percentage of immobilized lipid present at each temperature and the separation between the outermost hyperfine lines for the fluid lipid component. Lipid immobilization was attributed to lipid-protein interactions, lipid-cell wall interactions, and temperature-induced lipid phase transitions to the gel-state. Temperature profiles were compared for both cold-hardened and nonhardened protoplasts, plasma membranes, and plasma membrane lipids, respectively. Although cold-hardening extended the range of lipid fluidity by 5 degrees C, it had no effect on lipid-protein interactions or activation energies of lipid mobility. Differences were found, however, between the temperature profiles for the different samples, suggesting that alterations in the plasma membrane occurred as a consequence of the isolation methods used.

Journal Article↗
PubMed2001

MR ventilation-perfusion imaging of human lung using oxygen-enhanced and arterial spin labeling techniques.

Magnetic resonance ventilation-perfusion (V/Q) imaging has been demonstrated using oxygen and arterial spin labeling techniques. Inhaled oxygen is used as a paramagnetic contrast agent in ventilation imaging using a multiple inversion recovery (MIR) approach. Pulmonary perfusion imaging is conducted using a flow-sensitive alternating inversion recovery with an extra radiofrequency pulse (FAIRER) technique. A half Fourier single-short turbo spin echo (HASTE) sequence is used for data acquisition in both techniques. V/Q imaging was performed in ten of the twenty volunteers, while either ventilation or perfusion was imaged in the other ten. This V/Q imaging scheme is completely noninvasive, does not involve ionized radiation, and shows promising potential for clinical use in the diagnosis of lung diseases such as pulmonary embolism.

Adult↗
PubMed2005

Explanation of spin-lattice relaxation rates of spin labels obtained with multifrequency saturation recovery EPR.

Electron paramagnetic resonance (EPR) pulsed saturation recovery (pSR) measurements of spin-lattice relaxation rates have been made on nitroxide-containing fatty acids embedded in lipid bilayers by Hyde and co-workers. The data have been collected for a number of spin-labeled fatty acids at several microwave spectrometer frequencies (from 2 to 35 GHz). We compare these spin-lattice relaxation rates to those predicted by the Redfield theory incorporating several mechanisms. The dominant relaxation mechanism at low spectrometer frequencies is the electron-nuclear dipolar (END) process, with spin rotation (SR), chemical shift anisotropy (CSA), and a generalized spin diffusion (GSD) mechanism all contributing. The use of a wide range of spectrometer frequencies makes clear that the dynamics cannot be modeled adequately by rigid-body isotropic rotational motion. The dynamics of rigid-body anisotropic rotational motion is sufficient to explain the experimental relaxation rates within the experimental error. More refined models of the motion could have been considered, and our analysis does not rule them out. However, the results demonstrate that measurements at only two suitably chosen spectrometer frequencies are sufficient to distinguish anisotropic from isotropic motion. The results presented demonstrate that the principal mechanisms responsible for anisotropically driven spin-lattice relaxation are well understood in the liquids regime.

Electron Spin Resonance Spectroscopy↗
PubMed1979

The study of nitroxide radical active esters as spin labels on muscle protein actin.

Nitroxide radical active esters were used to label the muscle protein actin and to study the orientation dependence of the EPR spectra of the labelled protein. It is concluded that the labels are located at least at two different sites of the protein with strong polar environment and different mobilities. The EPR spectrum of the strongly immobilized labels exhibits orientation dependence, the N--O bond axis of the spin labels is nearly perpendicular to the long axis of the F-actin threads, and the labels undergo a rapid rotational motion about an axis directed perpendicular to the filament axis. The application of the nitroxide radical active esters may be useful in the study of ordered systems as muscle or muscle protein systems.

Actins↗
PubMed2000

Atomic structures of two nitroxide spin labels complexed with human thrombin: comparison with solution studies.

Crystal structures of thrombin complexed with two spin labels called para-V, 4-(2,2,5,5-tetramethylpyrrolidine-1-oxyl)-p-(fluorosulfonyl) benzamidine, and meta-V, 3-(2,2,5,5-tetramethyl-pyrrolidine1-oxyl)-m-(fluorosulfonyl) benzamidine, have been completed at 2.0 and 3.0 A resolution, respectively. Previous electron spin resonance studies with these labels gave rise to a low-resolution "topography map" of thrombin's extended active site. These labels monitor two distinct areas of the thrombin active site: (1) an apolar binding site which manifests itself in an biphasic activation/inhibition effect on thrombin activity and (2) a region sensitive to alpha-thrombin autoproteolytic cleavage(s) to gamma-thrombin (Arg75-Tyr76 and/or Arg77A-Asn78, and Lys149E-Gly150, chymotrypsin numbering). Para-V was found to bind along the substrate binding cleft, while meta-V was found to bind both at the substrate primary specificity pocket and at a site which interacts with the gamma-cleavage loop. These studies reaffirm that accurate information may be gained from solution studies and indicates the complementarity of solid-state studies.

Binding Sites↗
PubMed1991

Mobility of the spin-labeled side chains of some novel antifolate inhibitors in their complexes with dihydrofolate reductase.

Four spin-labeled inhibitors of dihydrofolate reductase (DHFR) have been synthesized, each of which has the 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO) reporting group at a different distance from the 2,4-diaminopyrimidine moiety by which the inhibitors are anchored and oriented in the active site. Inhibitors in which the TEMPO group is attached by a short side chain are weakly bound to DHFR from bacteria (Streptococcus faecium and Lactobacillus casei), to the bovine enzyme and to recombinant human DHFR. However, binding is sufficiently tight, especially in the ternary complexes with NADPH, for recording of the EPR spectra of the bound ligands. The spectra indicate that when these inhibitors are bound to the enzyme the TEMPO group is highly immobilized with correlation time, tau c, 4-20ns. Inhibitors that have the reporter group attached to the glutamate moiety of methotrexate bind to all four DHFRs more tightly than the inhibitors with shorter side chains by factors of up to 10(6). However, in most complexes formed by the inhibitors with longer side chains immobilization of the TEMPO group is slight (tau c 0.2-4 ns). These results are in general agreement with predictions from X-ray crystallographic results including thermal factors but there are some unanticipated differences between some results for bacterial and eukaryotic enzymes. Three of the splin-labeled inhibitors would provide good probes for distance measurements in and around the active site of mammalian DHFR.

Animals↗
PubMed1981

Interaction of selectively spin-labeled 70S ribosomes with tRNAPhe. An electron paramagnetic resonance study.

70S ribosomes from Escherichia coli, selectively spin labeled on the SH groups of proteins S18, S12, S21, S17, and L27, were used to study the formation of the tertiary complex ribosome-poly(U)-tRNAPhe. Most of these ribosomal proteins are located in the region of binding of tRNA. The electron paramagnetic resonance observable structural change suggests a loosening of the ribosome structure upon binding of the tRNA molecule.

Ascorbic Acid↗
PubMedDate not supplied

[Characterization of dynamic behavior of side groups of globular proteins by spin-label, NMR and luminescence polarization methods].

In this work we suggest a quantitative estimation of a complicated motion of side groups of globular proteins. In the general case, three basic parameters determine the motion: (a) rotational correlation time of a side unit under study, or covalently bound spin label, or dye, (b) parameter S that reflects sterical restrictions for re-orientation of the given unit (these two parameters depending on the side-chain structure and its conformational change within the immediate dynamic protein surrounding whereas correlation times of side units on microviscosity in addition), (c) rotational correlation time of protein globule. These parameters can be measured by spin-label, NMR and fluorescence polarization techniques. An attempt to describe a complicated dynamic behaviour of side units of protein macromolecules with a single dynamics parameter--rotational correlation time--not only leads to a loss of part of information about the local structural dynamics of macromolecules but also can diminish the tau value.

Electron Spin Resonance Spectroscopy↗
PubMed1988

Action of vitamin E as antioxidant in phospholipid liposomal membranes as studied by spin label technique.

The action of alpha-tocopherol in the phosphatidylcholine liposomal membranes was studied by spin label technique in order to elucidate the role of vitamin E in the membranes. The fluidity of the liposomal membranes decreased with increasing concentrations of saturated phosphatidylcholine, cholesterol, and alpha-tocopherol. However, the physical effect of alpha-tocopherol was quite small at its physiological concentrations. The effects of structure of various kinds of chromanols on the membrane fluidity were studied. The fluidity of the liposomal membranes decreased with increasing length of the side chain at 2-position of 6-chromanols. This effect was smaller when the side chain at the 2-position did not possess branched methyl group. The effect of tocopherols on the membrane fluidity was larger in the inner part of the membranes. Ascorbic acid residing in the aqueous phase interacted with the nitroxide spin probe incorporated into the membranes and the rate was smaller as the nitroxide group was buried deeper in the phospholipid bilayer. The spin probes incorporated into liposomal membranes were consumed as the oxidation proceeded. alpha-Tocopherol suppressed both the oxidation of membranes and the decrease of spin probe, but when alpha-tocopherol was depleted, the spin probe decreased and the membranes became more rigid as the oxidation proceeded.

Antioxidants↗
PubMed2000

Simulation studies of high-field EPR spectra of spin-labeled lipids in membranes.

The high-field (i.e., 94 GHz) membrane EPR spectra of lipids spin labeled in their fatty acid chains have been simulated by using two limiting motional models. The aim was to identify the dynamic origin of the residual (g(xx) - g(yy)) anisotropy observed in the nonaxial EPR spectra of cholesterol-containing membranes. It is concluded that the residual spectral anisotropy arises from in-plane ordering of the lipid chains by cholesterol. The partial averaging of the (g(xx) - g(yy)) anisotropy was best described by restricted axial rotation with a frequency in the region of tau(-1)(R||) approximately 0.5-1 x 10(9) s(-1). Simulations for slower axial rotation of unrestricted amplitude produced less satisfactory fits. In phospholipid membranes not containing cholesterol, the nonaxial anisotropy is completely averaged in the fluid phase and substantially reduced even in the gel phase. The unrestricted axial rotation in the gel phase is of comparable frequency to that of the limited axial rotation in the liquid-ordered phase of membranes containing cholesterol. These results on in-plane ordering by cholesterol in the liquid-ordered phase could be significant for current proposals regarding domain formation in cellular membranes.

Algorithms↗