Correlation Time Tau Research Articles

Narrow carbonyl resonances in proton-diluted proteins facilitate NMR assignments in the solid-state

HNCO/HNCACO type correlation experiments are an alternative for assignment of backbone resonances in extensively deuterated proteins in the solid-state, given the fact that line widths on the order of 14-17 Hz are achieved in the carbonyl dimension without the need of high power decoupling. The achieved resolution demonstrates that MAS solid-state NMR on extensively deuterated proteins is able to compete with solution-state NMR spectroscopy if proteins are investigated with correlation times tau(c) that exceed 25 ns.

Electron Spin Resonance Studies of the Reorientational Motion of Ni(mnt)2−

Electron spin resonance studies of the planar bis(maleonitriledithiolato)nickel complex ion, Ni(mnt)(2)(-), have been carried out from the motional narrowing region to the glassy limit in a series of ethers: 2-methyltetrahydrofuran (MTHF), diethylene glycol dimethyl ether (diglyme), triethylene glycol dimethyl ether (triglyme), and tetraethylene glycol dimethyl ether (tetraglyme). Analyses of the spectra show that Ni(mnt)(2)(-) is reorienting a factor of 3 faster about its long in-plane axis in all of these solvents; i.e., axially symmetric rotational diffusion produces agreement between the experimental and calculated line widths with D(parallel)/D(perpendicular) = 3.0 +/- 0.2; D(parallel) and D(perpendicular) are the diffusion constants for reorientation about the long in-plane (parallel) and perpendicular axes, respectively. The temperature dependence of the reorientational correlation time tau(2)(0) = (6D(perpendicular))(-1) determined from the widths is in agreement with the modified Stokes-Einstein-Debye model; the results indicate that Ni(mnt)(2)(-) has relatively strong (but not associative) interactions with the ethers. The experimental values of tau(2)(0) and the solvents' viscosities, self-diffusion constants, and dielectric relaxation times are compared and found to have a common temperature dependence. The ESR data also are compared with values of tau(solv), the correlation time obtained when a fluorescent probe is excited and its emission is monitored as the nonequilibrium solvent distribution relaxes. tau(solv) and tau(2)(0) are found to have a common temperature dependence in MTHF, tetraglyme, and two other solvents (ethyl alcohol and 1-butanol) in which Ni(mnt)(2)(-) has been studied. The factors determining these transport properties are discussed.

The solvation of nitroxide radicals in ionic liquids studied by high-field EPR spectroscopy

Ionic liquids (ILs) feature a variety of properties that make them a unique class of solvents. To gain a better understanding of how ILs solvate compounds of different chemical structure, we used pulsed high-field electron paramagnetic resonance (EPR) spectroscopy at W-band (approximately 94 GHz) and continuous wave EPR at X-band (approximately 9.4 GHz) on three TEMPO-based spin probes with different substitutions at the 4-position: 4-R-2,2,6,6-tetramethylpiperidine-1-oxyl, with R = N(CH(3))(3)(+), Cat-1, R = COO(-), TEMPO-4-carboxylate, and R = OH, TEMPOL. The spin probes are dissolved in imidazolium based ILs with different alkyl chain lengths (-C(2)H(5), -C(4)H(9), -C(6)H(13)) and anions (BF(4)(-), PF(6)(-)) and also in molecular solvents (methanol, water-glycerol). X-Band EPR at RT shows that the reorientational motion of the charged spin probes in ILs is about fivefold slower than that of the TEMPOL. Moreover, anion variation from BF(4)(-) to PF(6)(-) in ILs most strongly slows down the rotational motion (as measured by the rotational correlation time tau(r)) of Cat-1, followed by TEMPOL, while tau(r) of TEMPO-4-carboxylate is least affected. The EPR parameters g(xx) and A(zz) (tensor elements of the g- and hyperfine tensor) are sensitive to environmental effects and are only fully resolved at the high field used in this study. Changes of g(xx) and A(zz) values of the Cat-1 in ILs and methanol are very small especially compared to that of TEMPO-4-carboxylate, indicating that Cat-1 is located in a polar region of the ILs resembling the situation in methanol. On the other hand, the g(xx) value of TEMPO-4-carboxylate is sensitive to the length of alkyl group which shows that TEMPO-4-carboxylate is close to the nonpolar region of ILs. The small differences in the chemical substitution of the spin probes used here are sufficient for the molecules to reside in different domains of different dielectric properties in ILs. Our combined results are in good agreement with a picture of a nanophase separation, in which the charged cations and anions form polar regions and the hydrophobic alkyl chains of the IL cations form non-polar regions.

Femtosecond polarization resolved spectroscopy: A tool for determination of the three-dimensional orientation of electronic transition dipole moments and identification of configurational isomers

A method is presented that combines femtosecond polarization resolved UV/visible pump-IR probe spectroscopy and density functional theory calculations in determining the three-dimensional orientation of an electronic transition dipole moment (tdm) within the molecular structure. The method is demonstrated on the approximately planar molecule coumarin 314 (C314) dissolved in acetonitrile, which can exist in two ground state configurations: the E- and the Z-isomer. Based on an exhaustive search analysis on polarization resolved measurement data for four different vibrational modes, it is concluded that C314 in acetonitrile is the E-isomer. The electronic tdm vector for the electronic S(0)-->S(1) transition is determined and the analysis shows that performing the procedure for four vibrational modes instead of the minimally required three reduces the 1sigma probability area from 2.34% to 2.24% of the solution space. Moreover, the fastest rotational correlation time tau(c) for the C314 E-isomer is determined to be 26+/-2 ps.

Phosphatidylcholine “Wobble” in Vesicles Assessed by High-Resolution 13C Field Cycling NMR Spectroscopy

High resolution (13)C NMR field cycling (covering 11.7 down to 0.002 T) relaxation studies of the sn-2 carbonyl of phosphatidylcholines in vesicles provide a detailed look at the dynamics of this position of the phospholipid in vesicles. The spin-lattice relaxation rate, R(1), observed down to 0.05 T is the result of dipolar and CSA relaxation components characterized by a single correlation time tau(c), with a small contribution from a faster motion contributing to CSA relaxation. At lower fields, R(1) increases further with a correlation time consistent with vesicle tumbling. The tau(c) is particularly interesting since it is 2-3 times slower than what is observed for (31)P of the same phospholipid. However, cholesterol increases the tau(c) for both (31)P and (13)C sites to the same value, approximately 25 ns. These observations suggest faster local motion dominates the dipolar relaxation of the (31)P, while a slower rotation or wobble dominates the relaxation of the carbonyl carbon by the alpha-CH(2) group. The faster motion must be damped with the sterol present. As a general methodology, high resolution (13)C field cycling may be useful for quantifying dynamics in other complex systems as long as a (13)C label (without attached protons) can be introduced.

Coherence resonance induced by colored noise near Hopf bifurcation

Effects of colored noise near supercritical Hopf bifurcation, especially noise induced oscillation (NIO) and coherence resonance (CR), have been studied analytically in the Brusselator model, using the stochastic normal form method. Two types of colored noise are considered: one is the standard Gaussian colored noise generated by the Ornstein-Uhlenbeck (OU) process and the other is the so-called power-limited (PL) process. Depending on the noise intensity and noise type, it is found that the autocorrelation time, most probable radius and signal to noise ratio of the NIO may show nontrivial dependencies on the noise correlation time tau(c). Interestingly, for OU-type noise with intensity above a threshold, SNR is a bell-shaped function of tau(c), indicating enhancement of CR by noise correlation; and for PL-type noise, SNR may show double maxima when tau(c) is changed, demonstrating a new kind of multiresonance phenomenon. These theoretical predictions are well reproduced by numerical simulations.

Molecular Motions and Ion Diffusions of the Room-Temperature Ionic Liquid 1,2-Dimethyl-3-propylimidazolium Bis(trifluoromethylsulfonyl)amide (DMPImTFSA) Studied by 1H, 13C, and 19F NMR

The diffusive properties of an imidazolium room-temperature ionic liquid (RTIL), 1,2-dimethyl-3-propylimidazolium bis(trifluoromethylsulfonyl)amide (DMPImTFSA), are studied from the ionic conductivity and the ion diffusion coefficients measured by pulsed field gradient spin echo NMR. The temperature-dependent (1)H, (19)F, and (13)C NMR spin-lattice relaxation time T(1) values were observed, and the (1)H T(1) for DMPIm showed T(1) minima for various protons. According to the Bloemberger-Purcell-Pound (BPP) equation, the correlation time tau(c) values were directly calculated from (1)H NMR. By using the (1)H tau(c) values, an evaluation of the (13)C T(1) was attempted for the carbons having protons. The tau(c) estimated for molecular motions of DMPIm changes from 1.3 ns at 253 K to 72 ps at 353 K. The Stokes-Einstein-Debye (SED) model suggests that the tau(c) is too short for the overall molecular reorientation near room temperature. Consequently, the possibility of small-angle molecular rotation is proposed and tentative flip angles are calculated by using the translational diffusion coefficient, the bulk viscosity measured in this study, and the tau(c) obtained from (1)H T(1) data in the temperature range between 283 and 353 K. The flip amplitude increases with the temperature. DMPIm has isotropic reorientational motions with temperature-dependent amplitude, in addition to fast intramolecular motions such as methylene segmental motions, methyl rotational motion, and conformational exchange of the imidazolium ring. The existence of fast motions of TFSA is also shown. The translational diffusion of the ions is the slowest dynamic process in the present RTIL. Ab initio molecular orbital calculations are performed to understand the geometries of stable complexes of DMPIm(+) and TFSA(-), and the formation energies from the isolated ions are evaluated. The computed results are important for interpreting the (1)H T(1) behaviors observed for the imidazolium ring protons.

Dielectric Relaxation, Ion Conductivity, Solvent Rotation, and Solvation Dynamics in a Room-Temperature Ionic Liquid

Dielectric susceptibility and related conductivity of the neat ionic liquid 1-ethyl-3-methylimidazolium hexafluorophosphate (EMI+PF6-) are studied via molecular dynamics computer simulations. Both ion translations and reorientations contribute to dielectric relaxation, while their cross-correlation does not play any significant role. Interestingly, ion translational dynamics are found to enhance the static dielectric constant epsilon 0. The increment in epsilon 0 is attributed to rapid development of large anticorrelation in the autocorrelation function of the ionic current, i.e., hindered ion translations of strong librational character. One consequence of hindered translational dynamics is that the real part of conductivity has a maximum in the terahertz region and decreases with diminishing frequency. This in turn yields significant dielectric absorption in the far-IR region, consonant with recent terahertz time-domain spectroscopy measurements. Reorientational dynamics of cations show a marked deviation from diffusion. The well-known relation in the diffusion regime for reorientational correlation times tau R(l) proportional, variant [ l( l + 1)] (-1) fails completely for EMI+PF6-, where l is the order of Legendre polynomials used in the expansion of reorientational time correlation functions. It is found that dielectric continuum theory generally does not provide a reliable framework to describe solvation dynamics in EMI+PF6- even though the inclusion of ion conductivity in dielectric relaxation tends to improve the continuum description. This is ascribed mainly to electrostrictive effects absent in many continuum formulations.

Interspike interval statistics of a leaky integrate-and-fire neuron driven by Gaussian noise with large correlation times

We analytically investigate the interspike interval (ISI) density, the Fano factor, and the coefficient of variation of a leaky integrate-and-fire neuron model driven by exponentially correlated Gaussian noise with a large correlation time tau . We find a burstinglike behavior of the spike train, which is revealed by a dominant peak of the ISI density at small intraburst intervals and a slow power-law decay of long interburst intervals. The large, power-law distributed ISIs give rise to a coefficient of variation which diverges as square root [tau] . This leads to the paradoxical effect that ISI correlations, as expressed by the serial correlation coefficient, vanish for large correlation times. This is in contrast to findings of previous works on a simpler neuron model where the effect of noise correlations appeared in higher-order statistical measures.

Dynamic spin label study of the barstar-barnase complex

The dynamic spin label method was used to study protein-protein interactions in the model complex of the enzyme barnase (Bn) with its inhibitor barstar. The C40A mutant of barstar (Bs) containing a single cysteine residue was modified with two different spin labels varying in length and structure of a flexible linker. Each spin label was selectively bound to the Cys82 residue, located near the Bn-Bs contact site. The formation of the stable protein complex between Bn and spin labeled Bs was accompanied by a substantial restriction of spin label mobility, indicated by remarkable changes in the registered EPR spectra. Order parameter, S, as an estimate of rapid reorientation of spin label relative to protein molecule, was sharply increasing approaching 1. However, the rotational correlation time tau for spin-labeled Bs and its complex with Bn in solution corresponded precisely to their molecular weights. These data indicate that both Bs and its complex with Bn are rigid protein entities. Spin labels attached to Bs in close proximity to an interface of interaction with Bn, regardless of its structure, undergo significant restriction of mobility by the environment of the contact site of the two proteins. The results show that this approach can be used to investigate fusion proteins containing Bn or Bs.

Supramolecular Structure of Self‐Assembling Alamethicin Analog Studied by ESR and PELDOR

Three analogs of alamethicin F50/5, labelled with the TOAC (='2,2,6,6-tetramethylpiperidin-1-oxyl-4-amino-4-carboxylic acid') spin label at positions 1 (Alm1), 8 (Alm8), and 16 (Alm16), resp., were studied by Electron-Spin-Resonance (ESR) and Pulsed Electron-Electron Double-Resonance (PELDOR) techniques in solvents of different polarity to investigate the self-assembly of amphipathic helical peptides in membrane-mimicking environments. In polar solvents, alamethicin forms homogeneous solutions. In the weakly polar chloroform/toluene 1 : 1 mixture, however, this peptide forms aggregates that are detectable at 293 K by ESR in liquid solution, as well as by PELDOR in frozen, glassy solution at 77 K. In liquid solution, free alamethicin molecules and their aggregates show rotational-mobility correlation times tau(r) of 0.87 and 5.9 ns, resp. Based on these values and analysis of dipole-dipole interactions of the TOAC labels in the aggregates, as determined by PELDOR, the average number N of alamethicin molecules in the aggregates is estimated to be less than nine. A distance-distribution function between spin labels in the supramolecular aggregate was obtained. This function exhibits two maxima: a broad one at a distance of 3.0 nm, and a wide one at a distance of ca. 7 nm. A molecular-dynamics (MD)-based model of the aggregate, consisting of two parallel tetramers, each composed of four molecules arranged in a 'head-to-tail' fashion, is proposed, accounting for the observed distances and their distribution.

Application of Schrödinger Equation to Study the Tunnelling Dynamics of Proton Transfer in the Hydrogen Bond of 2,5-Dinitrobenzoic Acid: Proton T1, T1ρ, and Deuteron T1 Relaxation Methods

Temperature measurements of proton T1 (24.7 MHz), deuteron (deuterated hydroxyl group) T1 (55.2 MHz), and proton T1(rho) (B1 = 9 G) spin-lattice relaxation times of 2,5-dinitrobenzoic acid have been performed. An analysis of present experimental data together with previously published proton T1 (55.2 MHz) data has revealed the following molecular motions: proton/deuteron transfer in the hydrogen bond and two-site hopping of the whole dimer. It is shown that the proton-transfer dynamics are characterized by two correlation times tau(ov) and tau(tu), describing two fundamentally different motional processes, namely, thermally activated jumps over the barrier and tunneling through the barrier. The temperature dependence of 1/tau(tu) is the solution of Schrödinger's equation, which also yields the temperature T(tun), where begins the tunnel pathway for proton transfer. A new equation for the spectral density function of complex motion consisting of the three motions is derived. The third motion (two-site hopping of the whole dimer characterized by tau(lib) correlation time) is responsible for a proton T1(rho) minimum in high temperatures, just below the melting point. Such a minimum is not reached by T1 temperature dependencies. The minimum of T1(rho) assigned to the classical hopping of a hydrogen-bonded proton occurs in the same low-temperature regime in which the flattening of the temperature dependencies of T1 points to the dominance of incoherent tunneling. This experimental fact denies the known theories predicting the intermediate temperature regime where a smooth transition between classical and quantum tunneling dynamics is expected. The fit of the derived theoretical equations to the experimental data T1(rho) and T1 is satisfactory. The correlation times obtained for deuterons indicate deuteron-transfer dynamics much slower than proton-transfer dynamics. It is concluded that the classical proton transfer takes place over the whole temperature regime, while the incoherent tunneling occurs below 46.5 (hydrogen) or 87.2 K (deuterium) only.

Unexpected Aggregation of Neutral, Xylene‐Cored Dinuclear GdIII Chelates in Aqueous Solution

We have synthesized ditopic ligands L(1), L(2), and L(3) that contain two DO3A(3-) metal-chelating units with a xylene core as a noncoordinating linker (DO3A(3-) = 1,4,7,10-tetraazacyclododecane-1,4,7-triacetate; L(1) = 1,4-bis{[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]methyl}benzene; L(2) = 1,3-bis{[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]methyl}benzene; L(3) = 3,5-bis{[4,7,10-tris(carboxymethyl)-1,4,7,10-tetraazacyclododecane-1-yl]methyl}benzoic acid). Aqueous solutions of the dinuclear Gd(III) complexes formed with the three ligands have been investigated in a variable-temperature, multiple-field (17)O NMR and (1)H relaxivity study. The (17)O longitudinal relaxation rates measured for the [Gd(2)L(1-3)(H2O)(2)] complexes show strong field dependence (2.35-9.4 T), which unambiguously proves the presence of slowly tumbling entities in solution. The proton relaxivities of the complexes, which are unexpectedly high for their molecular weight, and in particular the relaxivity peaks observed at 40-50 MHz also constitute experimental evidences of slow rotational motion. This was explained in terms of self-aggregation related to hydrophobic interactions, pi stacking between the aromatic linkers, or possible hydrogen bonding between the chelates. The longitudinal (17)O relaxation rates of the [Gd(2)L(1-3)(H2O)(2)] complexes have been analysed with the Lipari-Szabo approach, leading to local rotational correlation times tau(1)(298) of 150-250 ps and global rotational correlation times tau(g)(298) of 1.6-3.4 ns (c(Gd): 20-50 mM), where tau(1)(298) is attributed to local motions of the Gd segments, while tau(g)(298) describes the overall motion of the aggregates. The aggregates can be partially disrupted by phosphate addition; however, at high concentrations phosphate interferes in the first coordination sphere by replacing the coordinated water. In contrast to the parent [Gd(DO3A)(H2O)(1.9)], which presents a hydration equilibrium between mono- and dihydrated species, a hydration number of q = 1 was established for the [Ln(2)L(1-3)(H2O)(2)] chelates by (17)O chemical shift measurements on Ln = Gd and UV/Vis spectrophotometry for Ln = Eu. The exchange rate of the coordinated water is higher for [Gd(2)L(1-3)(H2O)(2)] complexes k(ex)(298) = 7.5-12.0 x 10(6) s(-1)) than for [Gd(DOTA)(H2O)](-). The proton relaxivity of the [Gd(2)L(1-3)(H2O)(2)] complexes strongly decreases with increasing pH. This is related to the deprotonation of the inner-sphere water, which has also been characterized by pH potentiometry. The protonation constants determined for this process are logK(OH) = 9.50 and 10.37 for [Gd(2)L(1)(H2O)(2)] and [Gd(2)L(3)(H2O)(2)], respectively.

Analysis of NMR spin-lattice relaxation rates in cuprates

We investigate nuclear spin-lattice relaxation data in the normal state of optimally doped YBaCuO_7 by analyzing the contributions to the relaxation rate of the copper, planar oxygen and yttrium along the directions perpendicular to the applied field. In this new picture there is no contrasting temperature dependence of the copper and oxygen relaxation. We use the model of fluctuating fields to express the rates in terms of hyperfine interaction energies and an effective correlation time tau_eff characterizing the dynamics of the spin fluid. The former contain the effects of the antiferromagnetic static spin correlations, which cause the hyperfine field constants to be added coherently at low temperature and incoherently at high temperature. The degree of coherency is therefore controlled by the spin-spin correlations. The model is used to determine the temperature-dependent correlation lengths. The temperature-dependent effective correlation time is found to be made up of a linear and a constant contribution that can be related to scattering and spin fluctuations of localized moments respectively. The extrapolation of our calculation at higher temperature fits the data also very well at those temperatures. The underdoped compounds YBaCuO_6.63 and YBaCuO_8 are studied in the limit of the data available with some success by modifying the effective correlation time with a gap parameter. The copper data of the LaSrCuO series are then discussed in terms of the interplay between the two contributions to tau_eff.

Thermally activated escape rate for the Brownian motion of a fixed axis rotator in an asymmetrical double-well potential for all values of the dissipation

The Kramers theory of the escape rate of a Brownian particle from a potential well as extended by Mel'nikov and Meshkov is used to evaluate the relaxation times and the dynamic susceptibility for the rotational Brownian motion of fixed axis rotators in an asymmetric double-well potential. An expression for the escape rate valid for all values of the dissipation including the very low damping (VLD), very high damping (VHD), and crossover regimes is derived. It is shown that this expression provides a good asymptotic estimate of the inverse of the smallest nonvanishing eigenvalue lambda(1) of the underlying Fokker-Planck operator calculated by using the matrix-continued fraction method. For low barriers, where the Mel'nikov and Meshkov approach is not applicable, analytic equations for the correlation time tau( parallel) of the longitudinal dipole correlation function in the VLD and VHD limits are derived and a simple extrapolating equation valid for all values of the damping is proposed.

Transverse relaxation of cells labeled with magnetic nanoparticles

We describe the NMR relaxation properties of magnetically labeled cells. The cells are labeled with magnetic nanoparticles (SPIO, USPIO), which generate susceptibility contrast. The geometry of the labeled cells and the surrounding tissue is considered. We assume that the magnetic nanoparticles accumulate to form a magnetic core of radius RC inside the cell. The correlation time tau, which describes the motion of spins around this core, is analyzed. Using the strong collision approach, explicit expressions are derived for the transverse relaxation rate R2* for tissue containing labeled cells as a function of the core radius, the diffusion coefficient, and the concentration of the nanoparticles. The predictions of this model agree well with numerical simulations and experimental data.

Residue Specific Ribose and Nucleobase Dynamics of the cUUCGg RNA Tetraloop Motif by MNMR 13C Relaxation

The dynamics of the nucleobase and the ribose moieties in a 14-nt RNA cUUCGg hairpin-loop uniformly labeled with 13C and 15N were studied by 13C spin relaxation experiments. R1, R1rho and the 13C-[1H] steady-state NOE of C6 and C1' in pyrimidine and C8 and C1' in purine residues were obtained at 298 K. The relaxation data were analyzed by the model-free formalism to yield dynamic information on timescales of pico-, nano- and milli-seconds. An axially symmetric diffusion tensor with an overall rotational correlation time tau(c) of 2.31 +/- 0.13 ns and an axial ratio of 1.35 +/- 0.02 were determined. Both findings are in agreement with hydrodynamic calculations. For the nucleobase carbons, the validity of different reported 13C chemical shift anisotropy values (Stueber, D. and Grant, D. M., 2002 J. Am. Chem. Soc. 124, 10539-10551; Fiala et al., 2000 J. Biomol. NMR 16, 291-302; Sitkoff, D. and Case, D. A., 1998 Prog. NMR Spectroscopy 32, 165-190) is discussed. The resulting dynamics are in agreement with the structural features of the cUUCGg motif in that all residues are mostly rigid (0.82 < S2 < 0.96) in both the nucleobase and the ribose moiety except for the nucleobase of U7, which is protruding into solution (S2 = 0.76). In general, ribose mobility follows nucleobase dynamics, but is less pronounced. Nucleobase dynamics resulting from the analysis of 13C relaxation rates were found to be in agreement with 15N relaxation data derived dynamic information (Akke et al., 1997 RNA 3, 702-709).

Restricted orientational motion of nitroxides in molecular glasses: Direct estimation of the motional time scale basing on the comparative study of primary and stimulated electron spin echo decays

A comparative study of anisotropic relaxation in two-pulse primary and three-pulse stimulated electron spin echo decays provides a direct way to distinguish fast (correlation time tau(c)<10(-6) s) and slow (tau(c)>10(-6) s) motions. Anisotropic relaxation is detected as a difference of the decay rates for different resonance field positions in anisotropic electron paramagnetic resonance spectra. For fast motion anisotropic relaxation influences the primary echo decay and does not influence the stimulated echo decay. For slow motion it is seen in both two-pulse echo and three-pulse stimulated echo decays. For nitroxide spin probes dissolved in glassy glycerol only fast motion was found below 200 K. Increase of temperature above 200 K results in the appearance of slow motion. Its amplitude increases rapidly with temperature increase. While in glycerol glass slow motion appears above glass transition temperature T(g), in ethanol glass it is observable below T(g). The scenario of motional dynamics in glasses is proposed which involves the broadening of the correlation time distribution with increasing temperature.

Probing the Oligomeric State of Phospholamban Variants in Phospholipid Bilayers from Solid-State NMR Measurements of Rotational Diffusion Rates

Phospholamban (PLB) is a small transmembrane protein that regulates calcium transport across the sarcoplasmic reticulum (SR) of cardiac cells. PLB self-associates into pentamers within sodium dodecyl sulfate (SDS) micelles, but the oligomeric status of PLB in SR membranes is not known. This work has shown that a mutant of PLB, with all native cysteine residues replaced by alanine (Ala-PLB), runs as a monomer on SDS-PAGE gels, in agreement with previous studies [Karim et al. (2000) Biochemistry 39, 10892-10897]. By contrast, a peptide representing the transmembrane domain of the cysteine-free mutant (TM-Ala-PLB) coexists as pentamers, dimers, and monomers on gels. Solid-state NMR methods were used to examine the size and heterogeneity of Ala-PLB and TM-Ala-PLB labeled with (13)C and (2)H in the transmembrane domain and incorporated into dimyristoylphosphatidylcholine (DMPC) bilayers. Wide line (2)H NMR and (13)C cross-polarization magic-angle spinning (CP-MAS) NMR spectra of Ala-PLB and TM-Ala-PLB revealed two distinct species of each of the proteins in the membranes. In the case of Ala-PLB one species was present initially and a second species emerged after 12 h. Measurements of (1)H-(13)C dipolar couplings for the two species of Ala-PLB showed that the rotational diffusion of one species was relatively rapid, defined by a correlation time (tau(R)) of less than 10 micros, whereas the rotation of the other species was comparatively slow (tau(R) approximately 60 micros). These results suggest that although Ala-PLB runs as a monomer on gels, a mixture of different oligomeric forms of the protein, possibly monomers and pentamers, is present in DMPC bilayers. Caution must therefore be exercised in using SDS-PAGE to draw conclusions about the oligomeric state of PLB variants in lipid bilayers.

Study of the miscibility and segmental motion of STMAA-PBMA polymer blends and semi-interpenetrating polymer networks by an ESR spin probe method

Linear polymer blends and semi-interpenetrating polymer networks (IPNs) with controlled hydrogen bonding interactions based on poly(styrene-co-methacrylic acid) (STMAA) and poly(butyl methacrylate) (PBMA) were studied by an ESR spin probe method. The observed composite ESR spectra with fast- and slow-motion components in all the samples were ascribed to two-phase morphology. For linear blends, the temperatures T(a) corresponding to appearance of the fast motion, T(d) corresponding to the disappearance of slow motion, T(5mT) and the rotational correlation times tau(c) increased with increasing carboxylic acid content in STMAA. It was concluded that the degree of mixing of the blends was improved with increasing carboxylic acid content, owing to the enhanced hydrogen bonding interactions between the carboxylic acids in STMAA and the ester groups in PBMA. With respect to semi-IPN samples, there existed a competition in the microphase structure between the intercomponent hydrogen bonding interactions, which improved the miscibility of the samples and the intracomponent cross-linking, which might lead to phase separation in the systems with strong specific interactions. When the semi-IPN contained 29 mol% carboxylic acid, the temperatures T(d), T(5mT) and tau(c) reached their minimum values, which indicated that the sample reached its maximum miscibility.