基于NMR自旋弛豫技术的蛋白质动力学研究

蛋白质的三维结构在很多情况下不能很好地解释其在生理过程中的作用机制. 动力学研究能够获悉蛋白质在不同时间尺度下的内运动信息,建立起动态结构和生物功能的联系. 该文综述了通过NMR自旋弛豫技术研究蛋白质动力学的原理和方法：ps~ns的快运动分析主要采用约化谱密度函数映射和Modelfree方法;μs~ms的慢运动涉及化学/构象交换过程,常借助CPMG和R_1ρ弛豫色散手段. 基于NMR的蛋白质动力学研究,将蛋白质科学从三维空间结构推进到四维时空结构的新层面.     

生物分子结合水的结构与动力学研究进展

生物结合水在维护生物大分子的结构、稳定性以及调控动力学性质和生理功能等方面起着决定性的作用.从分子水平上理解生物结合水分子的结构与性质及其影响生物结构和功能的本质与规律,是揭示生物大分子生理功能机理的关键.目前生物结合水的结构与动力学相关研究尚处于初步阶段.本文从三个方面介绍当前生物结合水的相关研究及其进展:首先介绍结合水对蛋白质折叠、质子给予与迁移、配体结合与药物设计以及变构效应等生物结构和功能的影响;然后介绍生物分子周围的水分子结构研究情况;最后从时间尺度、动力学属性、生物分子与水分子之间的动力学耦合作用、蛋白质表面结合水次扩散运动等角度介绍生物分子水合动力学的研究进展,并归纳出一些目前尚待进一步解决的科学问题.     

129Xe and131Xe NMR of xenon on silica surfaces at 77 K

Little is known about¹²⁹Xe NMR spectral features and spin-lattice relaxation behavior, and the dynamics of xenon atoms, for xenon adsorbed on solid surfaces at cryogenic temperatures (≤77 K), where exchange with gas-phase atoms is not a significant complication. We report¹²⁹Xe NMR experiments at 9,4 T that provide such information for xenon adsorbed onto the hydroxylated surface of a number of microporous silica samples at 77 K. A convenient design for these cryogenic experiments is described. Dynamics of surface-adsorbed xenon atoms on the time scale of seconds can be observed by¹²⁹Xe NMR hole-burning experiments; much slower dynamics occurring over hours and days are evidenced from changes with time of the¹²⁹Xe NMR chemical shifts. The peak maxima occur in the region ca. 180–316 ppm, considerably downfield of¹²⁹Xe shifts previously reported on surfaces at higher temperatures, and closer to the shift of xenon bulk solid (316.4±1 ppm). The¹²⁹Xe spin-lattice relaxation timesT ₁ range over five orders of magnitude; possible explanations for both nonexponential relaxation behavior and extremely shortT ₁ values (35 ms) are discussed. Preliminary¹³¹Xe and¹H NMR results are presented, as well as a method for greatly increasing the sensitivity of¹²⁹Xe NMR detection at low temperatures by using closely-spaced trains of rf pulses.     

Double quantum filtered (1)H NMR spectroscopy enables quantification of lactate in muscle

In this study we address the question of quantification of muscle lactate using double quantum filtered (DQF) (1)H NMR spectroscopy where dipolar and scalar coupled spectra are acquired. For this, lactate content in muscle samples was independently determined using a conventional enzymatic assay and DQF, (1)H NMR spectroscopy. NMR quantification of lactate relied on comparison of muscle spectra with similarly acquired spectra of standard lactate solutions. Transverse relaxation, T(2), and dipolar coupling effects were investigated at two different orientations of muscle fibers relative to B(o) and at various lactate concentrations. In all cases, we found a biexponential T(2) decay of the lactate methyl signal with a long T(2) of 142 ms (+/-8 ms, n=24) and a short T(2) of 37 ms (+/-6 ms, n=24). Lactate content of muscle determined by NMR spectroscopy agreed with the results obtained from enzymatic assays of the same samples provided that T(2) effects as well as the presence of both scalar and dipolar coupling interactions of lactate in muscle were taken into account.     

固体核磁共振谱学研究层状双氢氧化物

层状双氢氧化物(LDHs)作为一类非常重要的无机超分子材料,已经被广泛应用于催化、离子交换、生命科学等众多领域．固体核磁共振谱学是研究层状双氢氧化
物局域结构和动态特征的一种强有力手段,提供了非常丰富的信息．特别是最近,借助固体核磁共振在探索层状双氢氧化物的结构方面取得了非常重要的进展（如：阳离子的有序性信息）．该文主要介绍了最近40 年来固体核磁共振研究层状双氢氧化物方面的重要进展．     

129Xe NMR study of Xe adsorption on multiwall carbon nanotubes

¹²⁹Xe NMR spectroscopy has been used to study the adsorption of Xe on multi-wall carbon nanotubes (MWCNT). The results obtained have shown the ¹²⁹Xe NMR ability to probe the intercrystalline (aggregate) and the inner porosity of CNT. In particular, the effects on porosity of tubes openings by hydrogen exposure and of ball milling were examined. Dramatic changes observed in the ¹²⁹Xe NMR spectra after moderate ball milling of MWCNTs were attributed to the destruction of the initial intercrystalline pore structure and to the Xe access inside the nanotubes. To examine the exchange dynamics the mixture of as-made and milled MWCNTs was studied with one- and two-dimensional (1D and 2D) ¹²⁹Xe NMR. The exchange between the interior of milled nanotubes and the aggregate pores of as-made MWCNTs was fast on the NMR acquisition time scale. The Xenon exchange between the interior of the as-made MWCNTs and the large aggregate pores occurred on a longer time scale of 10 ms, as was established by 2D ¹²⁹Xe NMR exchange spectroscopy. Variable temperature ¹²⁹Xe NMR data were also discussed and analyzed in terms of the fast exchange approximation.     

Role of protein-water hydrogen bond dynamics in the protein dynamical transition

The role of water in protein dynamics has been investigated using molecular dynamics simulations of crystals and a dehydrated powder. On the 100 ps time scale, the anharmonic and diffusive motions involved in the protein structural relaxation are correlated with the protein-water hydrogen bond dynamics. The complete structural relaxation of the protein requires relaxation of the hydrogen bond network via solvent translational displacement. Inhibiting the solvent translational mobility, and therefore the protein-water hydrogen bond dynamics, has an effect on the protein relaxation similar to dehydration.     

A Simple Approach to Analyzing Protein Side-Chain Dynamics fromC NMR Relaxation Data

A simple approach to deriving motional dynamics information of protein and peptide side chains by using¹³C NMR relaxation data is presented. By using linear approximation of internal rotational correlation functions, simple equations for relating side-chain conformation, bond rotational amplitudes, and rotational correlation coefficients with different NMR relaxation parameters have been obtained. Auto- and cross-correlation spectral densities are considered, and it is shown that proton-coupled¹³C NMR relaxation measurements allow detailed motional information to be obtained.     

Effects of various types of molecular dynamics on 1D and 2D (2)H NMR studied by random walk simulations

By carrying out random walk simulations we systematically study the effects of various types of complex molecular dynamics on (2)H NMR experiments in solids. More precisely, we calculate one-dimensional (1D) (2)H NMR spectra and the results of two dimensional (2D) (2)H NMR experiments in time domain, taking into account isotropic as well as highly restricted motions which involve rotational jumps about different finite angles. Although the dynamical models are chosen to mimic the primary and secondary relaxation in supercooled liquids and glasses, we do not intend to describe experimental results quantitatively but rather to show general effects appearing for complex reorientations. We carefully investigate whether 2D (2)H NMR in time domain, which was originally designed to measure correlation times of ultraslow motions (tau >/= 1 ms), can be used to obtain shorter tau, too. It is demonstrated that an extension of the time window to tau >/= 10 &mgr;s is possible when dealing with exponential relaxation, but that it will fail if there is a distribution of correlation times G(lgtau). Vice versa, we show that 1D (2)H NMR spectra, usually recorded to look at dynamics with tau in the microsecond regime, are also applicable for studying ultraslow motions provided that the loss of correlation is achieved step by step. Therefore, it is useful to carry out 1D and 2D NMR experiments simultaneously in order to reveal the mechanism of complex molecular motions. In addition, we demonstrate that highly restricted dynamics can be clearly observed in 1D spectra and in 2D NMR in time domain if long solid-echo delays and large evolution times are applied, respectively. Finally, unexpected observations are described which appear in the latter experiment when considering very broad distributions G(lgtau). Because of these effects, time scale and geometry of a considered motion cannot be extracted from a straightforward analysis of experimental results. Copyright 2000 Academic Press.     

Ultrafast α-like relaxation of a fragile glass-forming liquid measured using two-dimensional infrared spectroscopy

Ultrafast two-dimensional infrared (2D-IR) spectroscopy is used to study the picosecond dynamics of a vibrational probe molecule dissolved in a fragile glass former. The spectral dynamics are observed as the system is cooled to within a few degrees of the glass transition temperature (T(g)). We observe nonexponential relaxation of the frequency-frequency correlation function, similar to what has been reported for other dynamical correlation functions. In addition, we see evidence for α-like relaxation, typically associated with long-time, cooperative molecular motion, on the ultrafast time scale. The data suggests that the spectral dynamics are sensitive to cooperative motion occurring on time scales that are necessarily longer than the observation time.     

Scaling laws and memory effects in the dynamics of liquids and proteins

Recent progress in the numerical calculation of memory functions from molecular dynamics simulations allowed the gaining of deeper insight into the relaxation dynamics of liquids and proteins. The concept of memory functions goes back to the work of R. Zwanzig on the generalized Langevin equation, and it was the basis for the development of various dynamical models for liquids. In this article we present briefly a method for the numerical calculation of memory functions, which is then applied to study their scaling behavior in normal and fractional Brownian dynamics. It has been shown recently that the model of fractional Brownian dynamics constitutes effectively a link between protein dynamics on the nanosecond time scale, which is accessible to molecular dynamics simulations and thermal neutron scattering, and the much longer time scale of functional protein dynamics, which can be studied by fluorescence correlation spectroscopy. The text was submitted by the authors in English. Affiliated with the University of Orléans.     

核磁共振波谱在药物发现中的应用

核磁共振波谱通过检测组成有机化合物分子的原子核跃迁而得到反映核性质的参数以及周围化学环境对这些参数的影响规律. 这些参数的相关内容包含了极其详尽的有机化合物分子结构和分子间相互作用的信息,并构成了核磁共振结构解析和生物靶分子-配体相互作用研究的理论基础. 在生物医药研发领域内,科研院所和公司企业的研发工作者们一直在努力探索利用核磁共振波谱监测生物靶分子-配体相互作用作为药物发现工具的潜能. 本文旨在针对核磁共振波谱在药物发现过程中活性化合物筛选的最新研究进展进行综述.     

Correlation of primary and secondary relaxations in a supercooled liquid

The widespread assumption that primary and secondary relaxations in glass-forming materials are independent processes is scrutinized using spin-lattice relaxation weighted stimulated-echo spectroscopy. This nuclear magnetic resonance (NMR) technique is simultaneously sensitive to the dynamics on well-separated time scales. For the deeply supercooled liquid sorbitol, which exhibits a strong secondary relaxation, the primary relaxation (that is observable using NMR) can be modified by suppressing the contributions of those subensembles which are characterized by relatively slow secondary relaxations. This is clear evidence for a correlation between primary and secondary relaxation times. In the disordered crystal orthocarborane high-frequency processes are absent and consequently no such modifications could be achieved.     

Cooling overall spin temperature: protein NMR experiments optimized for longitudinal relaxation effects

In experiments performed on protonated proteins at high fields, 80% of the NMR spectrometer time is spent waiting for the ¹H atoms to recover their polarization after recording the free induction decay. Selective excitation of a fraction of the protons in a large molecule has previously been shown to lead to faster longitudinal relaxation for the selected protons [K. Pervushin, B. Vögeli, A. Eletsky, Longitudinal ¹H relaxation optimization in TROSY NMR spectroscopy, J. Am. Chem. Soc. 124 (2002) 12898–12902; P. Schanda, B. Brutscher, Very fast two-dimensional NMR spectroscopy for real-time investigation of dynamic events in proteins on the time scale of seconds, J. Am. Chem. Soc. 127 (2005) 8014–8015; H.S. Attreya, T. Szyperski, G-matrix Fourier transform NMR spectroscopy for complete protein resonance assignment, Proc. Natl. Acad. Sci. USA 101 (2004) 9642–9647]. The pool of non-selected protons acts as a “thermal bath” and spin-diffusion processes (“flip-flop” transitions) channel the excess energy from the excited pool to the non-selected protons in regions of the molecule where other relaxation processes can dissipate the excess energy. We present here a sensitivity enhanced HSQC sequence (COST-HSQC), based on one selective E-BURP pulse, which can be used on protonated ¹⁵N enriched proteins (with or without ¹³C isotopic enrichment). This experiment is compared to a gradient sensitivity enhanced HSQC with a water flip-back pulse (the water flip-back pulse quenches the spin diffusion between ¹H^N and ¹H^α spins). This experiment is shown to have significant advantages in some circumstances. Some observed limitations, namely sample overheating with short recovery delays and complex longitudinal relaxation behaviour are discussed and analysed.     

Dielectric and conductivity relaxation in mixtures of glycerol with LiCl

We report a thorough dielectric characterization of the α relaxation of glass-forming glycerol with varying additions of LiCl. Nine salt concentrations from 0.1 to 20mol% are investigated in a frequency range of 20Hz-3GHz and analyzed in the dielectric loss and modulus representation. Information on the dc conductivity, the dielectric relaxation time (from the loss) and the conductivity relaxation time (from the modulus) is provided. Overall, with increasing ion concentration, a transition from reorientationally to translationally dominated behavior is observed and the translational ion dynamics and the dipolar reorientational dynamics become successively coupled. This gives rise to the prospect that, by adding ions to dipolar glass formers, dielectric spectroscopy may directly couple to the translational degrees of freedom determining the glass transition, even in frequency regimes where usually strong decoupling is observed.     

Measurement of longitudinal relaxation times in crowded 1H NMR spectra using one- and two-dimensional maximum quantum (MAXY) NMR spectroscopy

Methods for measuring longitudinal relaxation times of protons in heavily overlapped ¹H NMR spectra are introduced and exemplified using a solution of cholesteryl acetate. The methods are based on 1-dimensional and 2-dimensional maximum quantum NMR spectroscopy (MAXY), which makes possible the selective detection of CH, CH₂ and CH₃ ¹H NMR resonances. A modification of the BIRD pulse sequence to achieve selective inversion of protons bonded to either ¹²C or ¹³C is given. The approach should find application in studies of molecular dynamics where isotopic enrichment is not possible and the level of available sample dictates the use of ¹H NMR spectroscopy.     

Sensitivity enhancement in (13)C solid-state NMR of protein microcrystals by use of paramagnetic metal ions for optimizing (1)H T(1) relaxation

We discuss a simple approach to enhance sensitivity for (13)C high-resolution solid-state NMR for proteins in microcrystals by reducing (1)H T(1) relaxation times with paramagnetic relaxation reagents. It was shown that (1)H T(1) values can be reduced from 0.4-0.8s to 60-70 ms for ubiquitin and lysozyme in D(2)O in the presence of 10 mM Cu(II)Na(2)EDTA without substantial degradation of the resolution in (13)C CPMAS spectra. Faster signal accumulation using the shorter (1)H T(1) attained by paramagnetic doping provided sensitivity enhancements of 1.4-2.9 for these proteins, reducing the experimental time for a given signal-to-noise ratio by a factor of 2.0-8.4. This approach presented here is likely to be applicable to various other proteins in order to enhance sensitivity in (13)C high-resolution solid-state NMR spectroscopy.     

Constrained modeling for spectroscopic measurement of bi-exponential spin-lattice relaxation of water in vivo

The (1)H NMR water signal from spectroscopic voxels localized in gray matter contains contributions from tissue and cerebral spinal fluid (CSF). A typically weak CSF signal at short echo times makes separating the tissue and CSF spin-lattice relaxation times (T(1)) difficult, often yielding poor precision in a bi-exponential relaxation model. Simulations show that reducing the variables in the T(1) model by using known signal intensity values significantly improves the precision of the T(1) measurement. The method was validated on studies on eight healthy subjects (four males and four females, mean age 21 +/- 2 years) through a total of twenty-four spectroscopic relaxation studies. Each study included both T(1) and spin-spin relaxation (T(2)) experiments. All volumes were localized along the Sylvian fissure using a stimulated echo localization technique with a mixing time of 10 ms. The T(2) experiment consisted of 16 stimulated echo acquisitions ranging from a minimum echo time (TE) of 20 ms to a maximum of 1000 ms, with a repetition time of 12 s. All T(1) experiments consisted of 16 stimulated echo acquisition, using a homospoil saturation recovery technique with a minimum recovery time of 50 ms and a maximum 12 s. The results of the T(2) measurements provided the signal intensity values used in the bi-exponential T(1) model. The mean T(1) values when the signal intensities were constrained by the T(2) results were 1055.4 ms +/- 7.4% for tissue and 5393.5 ms +/- 59% for CSF. When the signal intensities remained free variables in the model, the mean T(1) values were 1085 ms +/- 19.4% and 5038.8 ms +/- 113.0% for tissue and CSF, respectively. The resulting improvement in precision allows the water tissue T(1) value to be included in the spectroscopic characterization of brain tissue.     

Deuteron NMR relaxometry applied to confined liquid crystals

We discuss the capability of deuteron nuclear magnetic resonance (NMR) spectroscopy and relaxometry to reveal molecular ordering and dynamics in confined liquid crystals. The attention is focused on the high-temperature phase above the nematic-isotropic transition, which is — in the absence of the long-range orientational order — very suitable for the study of surface interactions. Deuteron NMR spectra and relaxation rates are presented for two representatives of confined liquidcrystal systems: 8CB in cylindrical cavities of Anopore membranes and 5CB with an embedded polymer network. A substantial increase in the transverse spin relaxation rate, stimulated by the surface-induced order in enclosures, has been observed. In cylindrical cavities, it exhibits a strong temperature dependence on approaching the phase transition, whereas in the polymer network dispersion it is temperature-independent. The increase of T ₂ ^?1 provides information on the effect of spatial constraints on molecular mobility and on the surface orientational order parameter. Using deuteron relaxometry, one can measure the degree of orientational order in the isotropic phase not only in cylindrical but also in spherical cavities and enclosures of irregular shape, where the standard approach based on quadrupolar splitting of the NMR spectrum fails.