P. Bánki

Magnetic tweezers for intracellular applications

We have designed and constructed a versatile magnetic tweezer primarily for intracellular investigations. The micromanipulator uses only two coils to simultaneously magnetize to saturation micron-size superparamagnetic particles and generate high magnitude constant field gradients over cellular dimensions. The apparatus resembles a miniaturized Faraday balance, an industrial device used to measure magnetic susceptibility. The device operates in both continuous and pulse modes. Due to its compact size, the tweezers can conveniently be mounted on the stage of an inverted microscope and used for intracellular manipulations. A built-in temperature control unit maintains the sample at physiological temperatures. The operation of the tweezers was tested by moving 1.28 μm diameter magnetic beads inside macrophages with forces near 500 pN.

Water fractions in normal and senile cataractous eye lenses studied by NMR

The state of water and water fractions in normal and senile cataractous eye lenses was studied by the NMR method. Combining NMR with vacuum dehydration provided additional information on multifractional samples. A new mathematical procedure is presented which separates the characteristic parameters of the different fractions and helps to determine the relaxation times and amounts of the fractions. The measurement accuracy enables separation of three different water fractions both in normal and in cataractous lenses.

Interfacial water at protein surfaces: Wide-line NMR and DSC characterization of hydration in ubiquitin solutions

Wide-line 1H-NMR and differential scanning calorimetry measurements were done in aqueous solutions and on lyophilized samples of human ubiquitin between -70 degrees C and +45 degrees C. The measured properties (size, thermal evolution, and wide-line NMR spectra) of the protein-water interfacial region are substantially different in the double-distilled and buffered-water solutions of ubiquitin. The characteristic transition in water mobility is identified as the melting of the nonfreezing/hydrate water. The amount of water in the low-temperature mobile fraction is 0.4 g/g protein for the pure water solution. The amount of mobile water is higher and its temperature dependence more pronounced for the buffered solution. The specific heat of the nonfreezing/hydrate water was evaluated using combined differential scanning calorimetry and NMR data. Considering the interfacial region as an independent phase, the values obtained are 5.0-5.8 J x g(-1) x K(-1), and the magnitudes are higher than that of pure/bulk water (4.2 J x g(-1) x K(-1)). This unexpected discrepancy can only be resolved in principle by assuming that hydrate water is in tight H-bond coupling with the protein matrix. The specific heat for the system composed of the protein molecule and its hydration water is 2.3 J x g(-1) x K(-1). It could be concluded that the protein ubiquitin and its hydrate layer behave as a highly interconnected single phase in a thermodynamic sense.

Diffusible and residual hydrogen in amorphous Ni(Cu)-Zr-H alloys

Abstract The partition of hydrogen into diffusible and residual parts was realized by pulse NMR spectroscopy, by gas chromatography and by prompt gamma activation analysis (PGAA). The total hydrogen content was determined by the two non-NMR methods and the diffusible (mobile) component by CPMG NMR pulse sequence. Results on amorphous Ni(Cu)–Zr–H systems of different compositions are shown. Partially crystallized samples were also studied as an extension. A method proposed by us directly gives the fractional population of hydrogen atoms in the free (mobile) state on the spin–spin relaxation time scale. On the other hand the least values of the residual hydrogen content correlate surprisingly well with the numbers of filled four Zr-type H-sites calculated by Batalla et al. [NATO ASI Ser. 136 (1985) 203] for 0.21-nm exclusion distance.

Hydration water/interfacial water in crystalline lens

Wide-line (1)H NMR signal intensity, spin-lattice and spin-spin relaxation rates and differential scanning calorimetry (DSC) measurements were done on avian (chicken and turkey) crystalline lenses between -70 degrees C and +45 degrees C to provide quantitative measures of protein hydration characteristic of the protein-water interfacial region. These measures are of paramount importance in understanding both the physiology of crystalline lens and its transitions to the cataractous pathological state characterized by the formation of opaque protein aggregates. Water mobility shows a characteristic transition at about -60 degrees C, which is identified as the melting of the interfacial/hydrate water. The amount of water in the low-temperature mobile fraction is about h = 0.4 g water/g protein, which equals the hydration required for protein activity. The amount of mobile water is temperature-independent up to about -10 degrees C, with a significant increase at higher temperatures below 0 degrees C. Above 0 degrees C, the relaxation processes can be described by a single (for spin-lattice) and by a triple (for spin-spin relaxation) exponential function. The spin-spin relaxation rate component of R(2) = 10-20 s(-1) and its dynamical parameters characterize the interfacial water at ambient or physiological temperatures. When considered an independent phase, the specific heat of the hydrate water obtained by a combination of DSC and NMR data in the temperature range -43 degrees C to -28 degrees C is higher than that of pure/bulk water. This discrepancy can only be resolved by assuming that the hydrate water is in strong thermodynamic coupling with the protein matrix. The specific heat for the system composed of the protein molecule and its hydration water is 4.6 +/- 0.3 J g(-1) K(-1). Thus, in a thermodynamic sense, crystalline protein and its hydrate layer behave as a highly-interconnected single phase.

Hydrogen occupancy, 1H NMR spectrum and second moment of ZrxNi1-x-H metallic glasses

Abstract The paper presents the results of an experimental investigation of rigid-lattice proton magnetic resonance (PMR) spectra and second moments in Ni 0.67 Zr 0.33 –H, Ni 0.50 Zr 0.50 –H, Ni 0.33 Zr 0.67 –H binary glassy alloy–hydrogen systems in the range of hydrogen-per-metal ( H / M ) ratio of 0.14≤ H / M ≤1.9. The line shape can be described by the empirical Harper-Barnes function with exponent values ranging from 1.45 to 2.95 in the whole hydrogen concentration range. The generally assumed Gaussian form is valid in a narrow H / M interval only. The second moment as a function of H / M can be fitted by a power function of an exponent 3/4. The exponent is independent of the composition of the alloy. We found that a simple lattice gas model based on a homogeneous hydrogen distribution contradicts the experimental results. Instead, a short-range order model has been proposed in which the hydrogen atoms are located in the centres of distorted tetrahedra of metal atoms, that is in the basic building blocks of the amorphous alloys. The Switendick criterion was also taken into consideration. We have shown that few proton first neighbours (0–2) and a small number of second neighbours (1–8), that is, a cluster of hydrogen is sufficient to account for the measured value of the second moment. Ternary Zr 0.5 Ni 0.5− x Cu x –H alloys have been investigated, too. The addition of Cu nuclei did not lead to the second moment enhancement expected for a homogeneous distribution of components.

High temperature 1H spin–spin relaxation in Zr–Ni–Cu–H amorphous alloys

Abstract 1 H spin–spin relaxation time ( T 2 ) and complementary hydrogen content, PMR spectrum width and spin–lattice relaxation time ( T 1 ) have been measured in Zr y (Ni 1− x Cu x ) 1− y –H ternary amorphous alloys of different hydrogen content at 0≤ x ≤33 at.% Cu and y =33, 50 and 67 at.% Zr concentrations using Carr–Purcell–Meiboom–Gill (CPMG), solid-echo and saturation recovery pulse sequences, respectively. At high hydrogen contents the T 2 measured by the slope of the CPMG echo train depends on both the Zr and Cu content, but is independent of the hydrogen content. The differences in the spin–spin relaxation behaviours can be attributed to the substantial change of correlation time and not to the change of activation energy or local fields. The measurements were made in the “motional narrowing” state, consequently our E a and τ ∞ quantities are averaged to the diffusion motion of protons taking part in this process. At small hydrogen contents T 2 depends on hydrogen content and the T 2 vs. 1/ T curves cannot be fitted by single Arrhenius plots. The hydrogen content measured by echo train has turned out to be systematically smaller than that measured by weight increase, demonstrating that not all the hydrogen takes part in the diffusion process.

PMR measurements on (Ni1 −xCux)0.5Zr0.5-Hy amorphous alloys

Abstract Spin-spin relaxation time ( T 2 ) and complementary hydrogen content (H/M), PMR spectrum width (FWHM) and spin lattice relaxation time ( T 1 ) have been measured in (Ni 1 − x Cu x ) 0.5 Zr 0.5 -H y ternary amorphous alloys at 0 ⩽ x ⩽ 29 at.% concentrations using Carr-Purcell-Meiboom-Gill (CPMG), solid-echo and saturation recovery pulse sequences respectively. The T 2 relaxation time measured by the slope of the CPMG echo train depends on both the hydrogen and Cu content. The dependence can be attributed to the change of correlation time and not to the change of activation energy or local field. The measurements were made in the ‘motional narrowing’ state; consequently our E a and τ ∞ quantities are averaged to the diffusional motion of protons taking part in this process. The extrapolated amplitude of the echo train measured on the alloy-hydrogen system and normalized to the same quantity in water, gives the value of proton magnetization in thermal equilibrium, that is the hydrogen content free from any perturbing effects. The hydrogen content measured by CPMG echo train has turned out to be systematically smaller than that measured by weight increase.

High temperature hydrogen diffusion in Zr0.33Ni0.67Hx amorphous alloys

Abstract The proton content in Zr 0.33 Ni 0.67 H x amorphous alloy was monitored by changes in electrical resistivity and with the extraplated spin echo amplitude measured by the Carr-Purcell-Meiboom-Gill (CPMG) nuclear magnetic resonance pulse sequence in the temperature interval 300 to 450 K. Both methods show that the hydrogen desorption processes can be approximately divided into two ranges. The first is a fast process which is at present not interpretable. In the second stage, the process is thermally activated, and the activation energy calcualted from the resistivity changes is equal to 0.32 ± 0.04 eV. The CPMG spin echo measurements also give the spin-spin relaxation time of diffusing protons and the activation energy of this process, 0.34 ± 0.02 eV, is exactly the same as extracted from the resistivity measurements.

Proton nuclear magnetic resonance and H-site occupancy in Zr0.5Ni0.5-yCuyHx metallic glasses

Abstract The aim of this paper is to reinvestigate the proton magnetic resonance spectra in Zr 0.5 Ni 0.5− y Cu y H x metallic glasses. Solid-echo pulse sequences were used to analyze the line shape. The measurements were made in the temperature range 2.2–300 K and at a frequency of 87.6 MHz. The reinvestigation of the line shape mostly at low temperatures was initiated after having realized that the splitting reported earlier on our samples was a consequence of the relatively long recovery time of our spectrometer. The present solid-echo measurements yielded simple spectra describable by a Harper-Barnes line shape close to gaussian.