Neutron Fibre Diffraction Research Articles

Solid–solvent molecular interactions observed in crystal structures of β-chitin complexes

Three β-chitin structures [anhydrous, di-hydrate, mono-ethylenediamine (EDA)] recently determined by synchrotron X-ray and neutron fiber diffraction were reviewed from the viewpoint of molecular interactions. Both water and EDA molecules interact with the chitin chains through multiple hydrogen bonds. When water complexes with chitin, the hydrogen bonding pattern rearranges with the replacement of an intrachain chitin hydrogen bond by a stronger hydrogen bond between chitin and water, with an associated reduction in the degrees of freedom; the water oxygen is a much stronger acceptor than the O5 ring atom. The behavior of hydrogen exchange by deuterium supports this interpretation. EDA-molecules change the conformation of hydroxymethyl group from gg to gt, accompanied by changes in hydrogen bonds due to the strong accepting ability of the EDA nitrogen atoms. Some important interactions are in common with experimental crystallographic results of cellulosic crystals and of molecular dynamics studies. These new insights into solid–solvent interactions are valuable in understanding molecular interactions in other polysaccharides-solvents system in solution or on surface.

Water in Crystalline Fibers of Dihydrate β-Chitin Results in Unexpected Absence of Intramolecular Hydrogen Bonding

The complete crystal structure (including hydrogen) of dihydrate β-chitin, a homopolymer of N-acetylglucosamine hydrate, was determined using high-resolution X-ray and neutron fiber diffraction data collected from bathophilous tubeworm Lamellibrachia satsuma. Two water molecules per N-acetylglucosamine residue are clearly localized in the structure and these participate in most of the hydrogen bonds. The conformation of the labile acetamide groups and hydroxymethyl groups are similar to those found in anhydrous β-chitin, but more relaxed. Unexpectedly, the intrachain O3-H…O5 hydrogen bond typically observed for crystalline β,1–4 glycans is absent, providing important insights into its relative importance and its relationship to solvation.

Direct Determination of the Hydrogen Bonding Arrangement in Anhydrous β-Chitin by Neutron Fiber Diffraction

The hydrogen bonding arrangement in anhydrous β-chitin, a homopolymer of N-acetylglucosamine, was directly determined by neutron fiber diffraction. Data were collected from a sample prepared from the bathophilous tubeworm Lamellibrachia satsuma in which all labile hydrogen atoms had been replaced by deuterium. Initial positions of deuterium atoms on hydroxyl and acetamide groups were directly located in Fourier maps synthesized using phases calculated from the X-ray structure and amplitudes measured from the neutron data. The hydrogen bond arrangement in the refined structure is in general agreement with predictions based on the X-ray structure: O3 donates a hydrogen bond to the O5 ring oxygen atom of a neighboring residue in the same chain; N2 and O6 donate hydrogen bonds to the same carbonyl oxygen O7 of an adjacent chain. The intramolecular O3···O5 hydrogen bond has the most energetically favorable geometry with a hydrogen to acceptor distance of 1.77 Å and a hydrogen bond angle of 171°.

Neutron fibre diffraction studies of amyloid using H2O/D2O isotopic replacement.

The first neutron fibre diffraction studies of an amyloid system are presented. The techniques used to prepare the large samples needed are described, as well as the procedures used to isotopically replace H2O in the sample by D2O. The results demonstrate the feasibility of this type of approach for the pursuit of novel structural analyses that will strongly complement X-ray fibre diffraction studies and probe aspects of amyloid structure that to date have remained obscure. The approach is demonstrated using an amyloid form of the peptide NSGAITIG, but is equally applicable for the study of other systems such as Alzheimer's Aβ peptide.

Looking at hydrogen bonds in cellulose

A series of cellulose crystal allomorphs has been studied using high-resolution X-ray and neutron fibre diffraction to locate the positions of H atoms involved in hydrogen bonding. One type of position was always clearly observed in the Fourier difference map (F(d)-F(h)), while the positions of other H atoms appeared to be less well established. Despite the high crystallinity of the chosen samples, neutron diffraction data favoured some hydrogen-bonding disorder in native cellulose. The presence of disorder and a comparison of hydrogen-bond geometries in different allomorphs suggests that although hydrogen bonding may not be the most important factor in the stabilization of cellulose I, it is essential for stabilizing cellulose III, which is the activated form, and preventing it from collapsing back to the more stable cellulose I.

Neutron diffraction measurements of skeletal muscle using the contrast variation technique: Analysis of the equatorial diffraction patterns

Among various methods for structural studies of biological macromolecules, neutron scattering and diffraction have a unique feature in that the contrast between the scattering length density of the molecules and that of the solvent can be varied easily by changing the D2O content in the solvent. This "contrast variation" technique enables one to obtain information on variations of scattering length density of the molecules of interest. Here, in order to explore the possibilities of the contrast variation technique in neutron fiber diffraction, neutron diffraction measurements of skeletal muscles were performed. The neutron fiber diffraction patterns from frog sartorius muscles were measured in various D2O concentrations in the relaxed state where no tension of muscle is produced, and in the rigor state where all myosin heads of the thick filaments bind tightly to actin in the thin filaments. It was shown that in both states, there were reflections having distinct contrast matching points, indicating a variation in the scattering length density of the protein regions in the unit cell of the muscle structure. Analysis of the equatorial reflections by the two-dimensional projected scattering length density map calculations by Fourier synthesis and model calculations provided the phase information of the equatorial reflections, with which the projected scattering length density maps of the unit cell of the hexagonal filament array in both states were calculated. The analysis showed that the scattering length density of the thick filament region was higher than that of the thin filament region, and that the scattering length density of the thick filament backbone changed as muscle went from the relaxed state into the rigor state.

Neutron fiber diffraction measurements of muscle using the contrast variation technique

Neutron fiber diffraction measurements of muscle using the contrast variation technique

The structure of celluloses

X-ray and neutron fiber diffraction has been used to study cellulose as it is converted from its naturally occurring crystal phase, cellulose I, to an activated crystal phase, cellulose IIII, by ammonia treatment. The detailed crystal structures determined for cellulose Iβ, an intermediate ammonia-cellulose I complex, and cellulose IIII, reveal a structural transition pathway: hydrogen bonded sheets of chains in cellulose Iβ slip with respect to each other to accommodate the penetrating ammonia guest molecules in the intermediate complex. On evaporation of ammonia from the intermediate complex, there is a relative small change in chain packing as an inter-sheet ammonia bridge is replaced by an inter-sheet hydrogen bond in cellulose IIII. When cellulose IIII is heated it converts back to cellulose Iβ. Both ammonia-cellulose I and cellulose IIII have extended chains of cooperative hydrogen bonds in relatively open crystal structures that may add to their susceptibility to rapid change.

Orientational Information of Troponin C within the Thin Filaments Obtained by Neutron Fiber Diffraction

In striated muscles contraction is regulated by the thin filament-based proteins, troponin consisting of three subunits (TnC, TnI, and TnT), and tropomyosin. Knowledge of in situ structures of these proteins is indispensable for elucidating this Ca 2+-sensitive regulatory mechanism. We employed neutron scattering to investigate the structure of TnC within the thin filament, and found that TnC assumes extended dumbbell-like structures and moves toward the filament axis by binding of Ca 2+. Here, in order to obtain more detailed in situ structural information of TnC, neutron fiber diffraction measurements were performed. Sols of native thin filaments and the thin filaments containing deuterated TnC were prepared in 2H 2O. The oriented samples were obtained by placing these sols sealed in quartz capillaries with a diameter of 3 mm in a magnetic field of 18 Tesla. Neutron fiber diffraction patterns were obtained from these oriented samples in the absence and presence of Ca 2+. The patterns obtained showed strong equatorial diffraction due to the thin filaments, 59 Å and 51 Å layer-lines due to actin, and meridional reflections due to Tn-complex. Analysis of the meridional reflections due to Tn-complex with aid of model calculation showed that the angle between the thin filament axis and the long axis of TnC was estimated to be 67(±7)° and 49(±17)°, in the absence and presence of Ca 2+, respectively, suggesting that TnC, which assumes orientations rather perpendicular to the filament axis in the absence of Ca 2+, tilts toward the filament axis and the orientational and positional disorder increases by binding Ca 2+. It also showed that the relative position of the TnC moved by about 22 Å by binding Ca 2+, and this apparent movement was concomitant with the movements of other Tn-subunits. This implies that by binding Ca 2+, significant structural rearrangements of Tn-subunits occur.

X-rays and neutrons for the study of DNA structure, hydration, and transitions

X-ray fibre diffraction methods have provided a major contribution to our understanding of a wide variety of biological polymers. However they are less effective for study of location of water and hydrogen atoms in these systems. Here neutron methods can provide vital information. The ability to deuterate biopolymers either throughout the entire molecule or in a more specific way adds a powerful dimension to work aimed at investigating hydration patterns or hydrogen positions and changes that occur during water-driven transitions. The purpose of this paper is to demonstrate how X-ray and neutron fibre diffraction methods can provide powerful probes of polymer structure when used in a genuinely complementary way. Two examples illustrating these issues are described. The first describes an X-ray fibre diffraction study of the A–B structural transition in natural DNA. The second describes preliminary results from the first neutron fibre diffraction study of a selectively labelled DNA polymer carried out on the instrument D19 at ILL using samples prepared in the ILL-EMBL Deuteration Laboratory.

Combined X-ray and neutron fibre diffraction studies of biological and synthetic polymers

The fibrous state is a natural one for polymer molecules which tend to assume regular helical conformations rather than the globular structures characteristic of many proteins. Fibre diffraction therefore has broad application to the study of a wide range of biological and synthetic polymers. The purpose of this paper is to illustrate the general scope of the method and in particular to demonstrate the impact of a combined approach involving both X-ray and neutron diffraction methods.While the flux of modern X-ray synchrotron radiation sources allows high quality datasets to be recorded with good resolution within a very short space of time, neutron studies can provide unique information through the ability to locate hydrogen or deuterium atoms that are often difficult or impossible to locate using X-ray methods. Furthermore, neutron fibre diffraction methods can, through the ability to selectively label specific parts of a structure, be used to highlight novel aspects of polymer structure that can not be studied using X-rays.Two examples are given. The first describes X-ray and neutron diffraction studies of conformational transitions in DNA. The second describes structural studies of the synthetic high-performance polymer poly(p-phenylene terephthalamide) (PPTA), known commercially as Kevlar® or Twaron®.

2P158 中性子繊維回折法による細いフィラメント中のトロポニンサブユニット配置の解析(筋肉(筋蛋白・収縮)))

2P158 中性子繊維回折法による細いフィラメント中のトロポニンサブユニット配置の解析(筋肉(筋蛋白・収縮)))

New Insights into the Structure of Poly(p-phenylene terephthalamide) from Neutron Fiber Diffraction Studies

New Insights into the Structure of Poly(<i>p</i>-phenylene terephthalamide) from Neutron Fiber Diffraction Studies

Cellulose IIII Crystal Structure and Hydrogen Bonding by Synchrotron X-ray and Neutron Fiber Diffraction

The crystal and molecular structure, together with the hydrogen-bonding system in ammonia-mercerized cellulose IIII, has been determined using synchrotron X-ray and neutron fiber diffraction data. The structure has a one-chain monoclinic unit cell with an asymmetric unit that contains only one glucosyl residue and with the hydroxymethyl group in the gt conformation. The hydrogen-bonding system is well-defined with no evidence of disorder. A bifurcated hydrogen bond links a donating secondary alcohol O3 atom to a ring O5 atom (major) and a primary alcohol O6 atom (minor) of an adjacent residue in the same chain. Two hydrogen bonds are present between neighboring chains, perpendicular to the chain direction. A detailed comparison of the crystal structure and hydrogen-bonding system reported here for cellulose IIII and those reported previously for the other cellulose polymorphs is given. The conformation of the chain in cellulose IIII has features similar to that of the center chain in the highly stable cel...

Water-DNA interactions as studied by X-ray and neutron fibre diffraction.

X-ray fibre-diffraction studies indicate a high degree of stereochemical specificity in interactions between water and the DNA double helix. Evidence for this comes from data that show that the molecular conformations assumed by DNA in fibres are highly reproducible and that the hydration-driven transitions between these conformations are fully reversible. These conformational transitions are induced by varying the relative humidity of the fibre environment and hence its water content. Further evidence for stereochemical specificity comes from the observed dependence of the conformation assumed on the ionic content of the fibre and the nucleotide sequence of the DNA. For some transitions, information on stereochemical pathways has come from real-time X-ray fibre diffraction using synchrotron radiation; information on the location of water with respect to the double helix for a number of DNA conformations has come from neutron fibre diffraction. This structural information from fibre-diffraction studies of DNA is complemented by information from X-ray single-crystal studies of oligonucleotides. If the biochemical processes involving DNA have evolved to exploit the structural features observed in DNA fibres and oligonucleotide single crystals, the challenges in developing alternatives to a water environment can be expected to be very severe.

D-118 Structure of Polymers Using Neutron and X-ray Fiber Diffraction—Invited

D-118 Structure of Polymers Using Neutron and X-ray Fiber Diffraction—<i>Invited</i>

1P179 細いフィラメントの中性子繊維回折(筋肉 : 筋蛋白・収縮)

1P179 細いフィラメントの中性子繊維回折(筋肉 : 筋蛋白・収縮)

Crystal structure and hydrogen bonding system in cellulose I(alpha) from synchrotron X-ray and neutron fiber diffraction.

The crystal and molecular structure, together with the hydrogen-bonding system in cellulose I(alpha), has been determined using atomic-resolution synchrotron and neutron diffraction data recorded from oriented fibrous samples prepared by aligning cellulose microcrystals from the cell wall of the freshwater alga Glaucocystis nostochinearum. The X-ray data were used to determine the C and O atom positions. The resulting structure is a one-chain triclinic unit cell with all glucosyl linkages and hydroxymethyl groups (tg) identical. However, adjacent sugar rings alternate in conformation giving the chain a cellobiosyl repeat. The chains organize in sheets packed in a "parallel-up" fashion. The positions of hydrogen atoms involved in hydrogen-bonding were determined from a Fourier-difference analysis using neutron diffraction data collected from hydrogenated and deuterated samples. The differences between the structure and hydrogen-bonding reported here for cellulose I(alpha) and previously for cellulose I(beta) provide potential explanations for the solid-state conversion of I(alpha) --> I(beta) and for the occurrence of two crystal phases in naturally occurring cellulose.

Crystal structure and hydrogen-bonding system in cellulose Ibeta from synchrotron X-ray and neutron fiber diffraction.

The crystal and molecular structure together with the hydrogen-bonding system in cellulose Ibeta has been determined using synchrotron and neutron diffraction data recorded from oriented fibrous samples prepared by aligning cellulose microcrystals from tunicin. These samples diffracted both synchrotron X-rays and neutrons to better than 1A resolution (>300 unique reflections; P2(1)). The X-ray data were used to determine the C and O atom positions. The resulting structure consisted of two parallel chains having slightly different conformations and organized in sheets packed in a "parallel-up" fashion, with all hydroxymethyl groups adopting the tg conformation. The positions of hydrogen atoms involved in hydrogen-bonding were determined from a Fourier-difference analysis using neutron diffraction data collected from hydrogenated and deuterated samples. The hydrogen atoms involved in the intramolecular O3...O5 hydrogen bonds have well-defined positions, whereas those corresponding to O2 and O6 covered a wider volume, indicative of multiple geometry with partial occupation. The observation of this disorder substantiates a recent infrared analysis and indicates that, despite their high crystallinity, crystals of cellulose Ibeta have an inherent disorganization of the intermolecular H-bond network that maintains the cellulose chains in sheets.

1P072中性子回折によるカエル骨格筋構造の解析

1P072中性子回折によるカエル骨格筋構造の解析