Adiabatic TOBSY in rotating solids

A MAS solid state NMR approach for achieving efficient scalar coupling mediated through-bond (13)C chemical shift correlations of the aliphatic carbons in uniformly labelled peptides/proteins is described. The method involves the application of a continuous train of adiabatic inversion pulses, as in the adiabatic TOCSY experiments carried out in solution state NMR studies. While rotor synchronised application of adiabatic inversion pulses leads to dipolar correlations, it is shown here via numerical simulations and experimental measurements that asynchronous application of adiabatic pulses can facilitate the mapping of through-bond connectivities. The method employs a suitable phasing scheme for generating the desired isotropic mixing Hamiltonian and requires moderate (13)C RF field strength only.     

Adiabatic Heteronuclear Decoupling in Rotating Solids

In magic angle spinning solid state NMR experiments the potential of heteronuclear (1)H decoupling employing a continuous train of adiabatic inversion pulses has been assessed via numerical simulations and experimental measurements. It is shown that, with a (1)H RF field strength of approximately 100 kHz that is typically available in MAS NMR probes, it is possible to achieve efficient adiabatic (1)H decoupling at low magic angle spinning frequencies. It is pointed out that in the presence of H (1) inhomogeneities it will be advantageous to employ adiabatic decoupling in MAS solid state NMR experiments.     

Tailoring broadband inversion pulses for MAS Solid state NMR

A simple approach is demonstrated for designing optimised broadband inversion pulses for MAS solid state NMR studies of biological systems. The method involves a two step numerical optimisation procedure and takes into account experimental requirements such as the pulse length, resonance offset range and extent of H₁ inhomogeneity compensation needed. A simulated annealing protocol is used initially to find appropriate values for the parameters that define the well known tanh/tan adiabatic pulse such that a satisfactory spin inversion is achieved with minimum RF field strength. This information is then used in the subsequent stage of refinement where the RF pulse characteristics are further tailored via a local optimisation procedure without imposing any restrictions on the amplitude and frequency modulation profiles. We demonstrate that this approach constitutes a generally applicable tool for obtaining pulses with good inversion characteristics. At moderate MAS frequencies the efficacy of the method is experimentally demonstrated for generating double-quantum NMR spectra via the zero-quantum dipolar recoupling scheme RFDR.     

Solid state NMR at high magic angle spinning frequencies: Dipolar chemical shift correlation with adiabatic inversion pulse based RF pulse schemes

The efficacy of hetero- and homonuclear dipolar recoupling employing tanh/tan adiabatic inversion pulse based RF pulse schemes has been examined at high magic angle spinning (MAS) frequencies via numerical simulations and experimental measurements. An approach for minimising the recoupling RF power level is presented, taking into consideration the spinning speed, the range of resonance offsets and H₁ inhomogeneities and the available RF field strength. This involves the tailoring of the frequency and amplitude modulation profiles of the inversion pulses. The applicability of tanh/tan pulse based dipolar recoupling schemes to spinning speed regimes where the performance with conventional rectangular pulses may not be satisfactory is demonstrated.     

Broadband RFDR with adiabatic inversion pulses

With a view to obtain ¹³C chemical shift correlation spectra of uniformly labelled peptides/proteins at high magnetic fields and high magic angle spinning frequencies (_r/2 20 kHz), the efficacy of RFDR with adiabatic inversion pulses has been assessed via numerical simulations and experimental measurements employing different adiabatic pulse phasing schemes, shapes and durations. It is demonstrated that homonuclear dipolar recoupling with superior performance under resonance offset and H ₁ inhomogeneity effects and without strong dependence on the ¹³C chemical shift differences can be achieved with adiabatic pulses. It is shown that ¹³C chemical shift correlation spectra in the entire range of carbon chemical shifts can be obtained efficiently with short adiabatic inversion pulses. In situations where correlation spectra of only the aliphatic region are required, the possibility for minimising the interference between the recoupling and decoupling RF fields with long adiabatic pulses, at low recoupling power levels and without compromising the broadband RFDR characteristics, is also indicated.     

Broadband homonuclear chemical shift correlation at high MAS frequencies: a study of tanh/tan adiabatic RF pulse schemes without $${^{{\bf 1}}\hbox{{\bf H}}}$$ decoupling during mixing

At high magic angle spinning (MAS) frequencies the potential of tanh/tan adiabatic RF pulse schemes for ¹³C chemical shift correlation without ¹H decoupling during mixing has been evaluated. It is shown via numerical simulations that a continuous train of adiabatic ¹³C inversion pulses applied at high RF field strengths leads to efficient broadband heteronuclear decoupling. It is demonstrated that this can be exploited effectively for generating through-bond and through-space, including double-quantum, correlation spectra of biological systems at high magnetic fields and spinning speeds with no ¹H decoupling applied during the mixing period. Experiments carried out on a polycrystalline sample of histidine clearly suggest that an improved signal to noise ratio can be realised by eliminating ¹H decoupling during mixing.     

Chemical shift correlation via RFDR: elimination of resonance offset effects

It is shown that it is possible to effectively execute RFDR experiments with adiabatic inversion pulses and obtain resonance offset compensation that is superior to what can be achieved by conventional rectangular pulses. Employing 40-s tanh/tan adiabatic pulses at a power level of 38 kHz and a spinning speed of 12 kHz it is demonstrated that the range of resonance offset compensation achieved is sufficient to generate, via a single experiment, homonuclear chemical shift correlation spectra in the entire ¹³C chemical shift range in peptides/proteins at the currently available field strengths.     

Adiabatic TOCSY for C,C and H,H J-transfer

Adiabatic pulses have been widely used for broadband decoupling and spin inversion at high magnetic fields. In this paper we propose adiabatic pulses and supercycles that can be used at high magnetic fields like 800 or 900 MHz to obtain broadband TOCSY sequences with C,C or H,H J-transfer. The new mixing sequences are equal or even superior to the well known DIPSI-2,3 experiments with respect to bandwidth. They prove robust against pulse miscalibration and B₁ inhomogeneity and are therefore attractive for fully automated spectrometer environments. These adiabatic mixing sequences have been incorporated in a novel z-filter HCCH-TOCSY experiment.     

Homonuclear chemical shift correlation in rotating solids via RN<Superscript>ν</Superscript><Subscript><Emphasis Type="Italic">n</Emphasis></Subscript> symmetry-based adiabatic RF pulse schemes

The efficacy of RN_n symmetry-based adiabatic Zero-Quantum (ZQ) dipolar recoupling schemes for obtaining chemical shift correlation data at moderate magic angle spinning frequencies has been evaluated. RN_n sequences generally employ basic inversion elements that correspond to a net 180° rotation about the rotating frame x-axis. It is shown here via numerical simulations and experimental measurements that it is also possible to achieve efficient ZQ dipolar recoupling via RN_n schemes employing adiabatic pulses. Such an approach was successfully used for obtaining ¹3C chemical shift correlation spectra of a uniformly labelled sample of (CUG)₉7– a triplet repeat expansion RNA that has been implicated in the neuromuscular disease myotonic dystrophy. An analysis of the ¹3C sugar carbon chemical shifts suggests, in agreement with our recent ¹5N MAS-NMR studies, that this RNA adopts an A-helical conformation.     

How Accurate Is MOLLI T1 Mapping In Vivo? Validation by Spin Echo Methods

T₁ mapping is a promising quantitative tool for assessing diffuse cardiomyopathies. The purpose of this study is to quantify in vivo accuracy of the Modified Look-Locker Inversion Recovery (MOLLI) cardiac T₁ mapping sequence against the spin echo gold standard, which has not been done previously. T₁ accuracy of MOLLI was determined by comparing with the gold standard inversion recovery spin echo sequence in the calf muscle, and with a rapid inversion recovery fast spin echo sequence in the heart. T₁ values were obtained with both conventional MOLLI fitting and MOLLI fitting with inversion efficiency correction. In the calf (n = 6), conventional MOLLI fitting produced inconsistent T₁ values with error ranging from 8.0% at 90° to 17.3% at 30°. Modified MOLLI fitting with inversion efficiency correction improved error to under 7.4% at all flip angles. In the heart (n = 5), modified MOLLI fitting with inversion correction reduced T₁ error to 5.5% from 14.0% by conventional MOLLI fitting. This study shows that conventional MOLLI fitting can lead to significant in vivo T₁ errors when not accounting for the lower adiabatic inversion efficiency often experienced in vivo.     

Mathematical treatment of adiabatic fast passage pulses for the computation of nuclear spin relaxation rates in proteins with conformational exchange

Although originally designed for broadband inversion and decoupling in NMR spectroscopy, recent methodological developments have introduced adiabatic fast passage (AFP) pulses into the field of protein dynamics. AFP pulses employ a frequency sweep, and have not only superior inversion properties with respect to offset effects, but they are also easily implemented into a pulse sequence. As magnetization is dragged from the +z to the -z direction, Larmor precession is impeded since magnetization becomes spin-locked, which is a potentially useful feature for the investigation of microsecond to millisecond dynamics. A major drawback of these pulses as theoretical prediction is concerned, however, results from their time-dependent offset: simulations of spin density matrices under the influence of a time-dependent Hamiltonian with non-commuting elements are costly in terms of computational time, rendering data analysis impracticable. In this paper we suggest several ways to reduce the computational time without compromising accuracy with respect to effects such as cross-correlated relaxation and modulation of the chemical shift.     

REDOR: an assessment of the efficacy of dipolar recoupling with adiabatic inversion pulses

A numerical assessment of the efficacy of REDOR recoupling of heteronuclear dipolar interactions employing adiabatic dephasing pulses has been carried out by considering an isolated dipolar coupled spin 1/2 I-S system. At moderate magic angle spinning frequencies in the range of 3–6 kHz and when the CSA of the dephased spins is large, it is shown that efficient broadband heteronuclear dipolar recoupling and reliable distance estimates can be achieved even under conditions where a significant fraction of the rotor period is occupied by the adiabatic pulse. The efficacy of REDOR with adiabatic inversion pulses has been demonstrated experimentally in two model ¹⁵N-¹³C spin systems, (¹³C,¹⁵N) Aib-(¹⁵N) Aib-NH₂ (Aib = -aminoisobutyric acid) and (1-¹³C,¹⁵N) glycine.     

Topologically fixed SecG is fully functional

It has been proposed that the bitopic membrane protein SecG undergoes topology inversion during translocation of (pre)proteins via SecYEG. Here we show that SecG covalently cross-linked to SecY cannot invert its topology while remaining fully functional in protein translocation. Our results strongly disfavor topology inversion of SecG during protein translocation.     

D型氨基酸氧化酶活性对于D-硝基精氨酸手性转化的影响

D-硝基精氨酸(D-NNA)可在大鼠体内发生手性转化生成其L型异构体,即L-NNA,后者可抑制一氧化氮合酶活性,减少一氧化氮生成,升高动脉血压.研究了D型氨基酸氧化酶(DAAO)在D-NNA手性转化中的作用及DAAO对不同(包括已报道在体内可发生手型转化的)D型氨基酸的选择活性.体内实验显示,DAAO的选择性抑制剂苯甲酸钠(400mg/kg)或肌酐(400mg/kg)均可在不同程度上抑制D-NNA升压作用,进一步研究发现,肾脏或肝脏DAAO酶液在外加DAAO后可提高D-NNA的手性转化约2倍,表明DAAO对于D-NNA在体内的手性转化是必需的.DAAO酶液对可在体内发生手性转化且转化率相似(30%～50%)的D型氨基酸(D-Phe,D-Leu和D-NNA)的选择性表现出显著差异(Kcat/Km相差可达约15倍左右),这从另一方面表明体内D-硝基精氨酸氧化是其发生手性转化的前提条件但非决定因素.     

Solid-phase synthesis of oligonucleotides containing a bipyridine ligand at the 3'-3' inversion of polarity site

The preparation of a solid support useful for the synthesis of oligonucleotides with a 3'-3' inversion of polarity, via a linker containing a chelating molecule, namely 2,2'-bipyridine, is described.     

Inversion of chiral α-methylbenzylamine

More effective use of optical resolution processes can be obtained by increasing the overall yields after development of methods for inversion of the chiral centre of the unwanted isomer. The configuration of some optically active amines can be inverted in a three-step synthesis via the N,N-ditosylimides and a subsequent nucleophilic substitution by the azide ion. The azide product is reduced by hydrogenolysis. Low stereoselectivity caused by racemization to some extent was at first observed for the inversion of the benzylic substrate, (S)-α-methylbenzylamine ( 5a ). However, modified reaction conditions allowed increased stereoselectivity, a more rapid and almost complete inversion of this substate as well. © 1994 Wiley-Liss, Inc.     

New enzymes for biotransformations

Several novel bioprocesses that have little or no counterpart in traditional methodology have recently been reported. The stereoselective and enantioselective hydrolysis of sec-alkyl sulfate esters by alkyl sulfatases proceeds with inversion of configuration and furnishes a homochiral product mixture. Haloalcohol dehalogenases were shown to accept various non-natural nucleophiles, such as azide, cyanide and nitrite for the asymmetric opening of epoxides giving rise to the corresponding azido-, cyano-, and nitro-alcohols as non-natural products. Asymmetric carbon-carbon bond formation via the acyloin- and benzoin-reaction was successfully catalyzed in water by novel lyases, such as benzoylformate decarboxylase and benzaldehyde lyase. New methods for the production of chiral nonracemic alpha-L-amino acids and amines were recently reported. Enantioselective stereoinversion of racemic alpha-aryl- and alpha-aryloxycarboxylic acids via epimerase-catalyzed inversion led to a single stereoisomeric product from the racemate.     

Inversion frequencies and phenotypic effects are modulated by the environment: insights from a reciprocal transplant study in <Emphasis Type="Italic">Coelopa frigida</Emphasis>

Understanding how environmental variation drives phenotypic diversification within species is a major objective in evolutionary biology. The seaweed fly Coelopa frigida provides an excellent model for the study of genetically driven phenotypes because it carries an α/β inversion polymorphism that affects body size. Coelopa frigida inhabits highly variable beds of decomposing seaweed on the coast in Scandinavia thus providing a suitable test ground to investigate the genetic effects of substrate on both the frequency of the inversion (directional selection) and on the phenotype (genotype?×?environment effects). Here we use a reciprocal transplant experiment to test the effect of the α/β inversion on body size traits and development time across four suitable natural breeding substrates from the clinal distribution. We show that while development time is unaffected by G?×?E effects, both the frequency of the inversion and the relative phenotypic effects of the inversion on body size differ between population?×?substrate combinations. This indicates that the environment modulates the fitness as well as the phenotypic effects of the inversion karyotypes. It further suggests that the inversion may have accumulated qualitatively different mutations in different populations that interact with the environment. Together our results are consistent with the idea that the inversion in C. frigida likely evolves via a combination of local mutation, G?×?E effects, and differential fitness of inversion karyotypes in heterogeneous environments.     

Synthesis of 2,3:4,6-di-O-isopropylidene-D-allopyranose from D-glucose

2,3:4,6-Di-O-isopropylidene-d-allopyranose can be conveniently prepared from d-glucose via a synthetic sequence, which includes Mitsunobu inversion at O-3, di-O-isopropylidenation of phenyl-1-thio-d-alloside and anomeric deprotection on treatment with NBS/CaCO3.     

Adiabatic solution of the Ohmic cable equation. General theory, homogeneous cylindrical fiber]

An adiabatic solution of the Ohmic cable equation is suggested, which reduces the non-stationary equation to a stationary form. The adiabatic length constant of the stationary equation is time-dependent. The adiabatic solutions for the boundary conditions that change in time linearly and exponentially were studied. In the latter case, the adiabatic length constant does not depend on time though it differs from the usual length constant. The cable input characteristics of exact and adiabatic solutions were compared in the cases of the voltage- and current-clamp, and electric field stimulation. The adiabatic and exact solutions are identical for the rising exponential stimuli. For the falling exponential stimuli, the adiabatic solution determines the exact asymptotic solution if the stimulus decays slower than the relaxation of initial conditions. It is propose to use linear and exponential ramp stimulation in electrotonic measurements.