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Biomedical subjects

Christian Riekel

Publications and source records attributed to Christian Riekel.

16 recordsLinked to original sources
PubMed2005

Morphological and structural aspects of the giant starch granules from Phajus grandifolius.

The morphology and structure of giant starch granules from the pseudo-bulbs of Phajus grandifolius were investigated, using a number of microscopy techniques together with synchrotron radiation microdiffration analysis. Most of the granules, which had sizes between 100 and 200 microm, occurred as ogival particles with the hilum or proximal end located at the apex of the granules. A small percentage of granules held a protuberance extending orthogonally to the underlying parent granule. Growth rings were observed in all granules: strongly curved close to the hilum, but planar toward the distal end of the granules or in the protuberances. Specific mechanical disruption followed by enzymatic digestion revealed the susceptibility of the disorganized parts of the growth rings, which were preferentially carved away during the digestion, leaving behind the better-organized domains. Microdiffraction analysis achieved with synchrotron radiation revealed the crystalline features of the granules and provided orientation maps of the amylopectin molecules in the various parts of the granules. In simple ogival granules the amylopectin molecules were uniformly oriented with their axes running from the hilum toward the distal end of the granule. In granules with a protuberance, the axes of the amylopectin molecules kept their direction in the parent granule, but took an orthogonal direction in the protuberance. The occurrence of these morphological and structural features is tentatively correlated with the mode of growth of these granules.

Cytoplasmic Granules↗
PubMed2005

microGISAXS and protein nanotemplate crystallization: methods and instrumentation.

Microbeam grazing-incidence small-angle X-ray scattering (microGISAXS) has been used and the technique has been improved in order to investigate protein nucleation and crystal growth, assisted by a protein nanotemplate. The aim is to understand the protein nanotemplate method in detail, as this method has been proved capable of accelerating and increasing crystal size and quality as well as inducing crystallization of proteins that are not crystallizable by classical methods. The nanotemplate experimental setup was used for drops containing growing lysozyme crystals at three different stages of growth.

Crystallography, X-Ray↗
PubMed2005

Structure and mechanical properties of carbon fibres: a review of recent microbeam diffraction studies with synchrotron radiation.

Hard X-ray beams with beam sizes in the sub-micrometre range are frequently available at third-generation synchrotron radiation sources, enabling new insights into the structure of materials at different levels of hierarchy by applying novel techniques such as scanning microbeam diffraction or scanning small-angle scattering. The high brilliance of the radiation from wiggler and undulator sources allows in situ experiments such as mechanical testing of single fibres to be performed, and even the combination of microbeam scanning with in situ testing is feasible. Three different experiments on single carbon fibres are presented: an in situ tensile test using a 10 microm collimated beam, a scanning experiment applying a 3 microm beam from a tapered glass capillary, and a scanning experiment with simultaneous in situ bending with a 0.1 microm beam provided by a waveguide structure.

Journal Article↗
PubMed2005

Studying morphological variations across single fibres using an X-ray waveguide.

Waveguide and glass capillary optics have been used to study morphological variations across single as-spun poly(p-phenylene benzobisoxazole) (PBO) fibres. The waveguide provides a beam with a submicrometre dimension perpendicular to the fibre axis. The glass capillary optics provide a beam of a few micrometres in diameter. By directly comparing results obtained with both optics it is possible to assess the influence of beam size on experimental observations. The study demonstrates that submicrometre beams are invaluable for detailed morphological studies across single fibres. The greater morphological averaging inherent to data collected using glass capillary optics reduces the variation between skin and core measurements, as well as obscuring other important features. Crystalline domain orientation, (200) crystal plane spacings and reflection intensities can be determined from data generated using a submicrometre beam. All parameters are found to vary significantly across the diameter of single PBO fibres. In terms of orientation, both skin and core regions exhibit a lower degree of crystalline domain orientation than an intermediate zone. In terms of (200) crystal plane spacing, the observed variations suggest residual stresses within the fibre structure following manufacture.

Journal Article↗
PubMed2005

Structure of the cross-beta spine of amyloid-like fibrils.

Numerous soluble proteins convert to insoluble amyloid-like fibrils that have common properties. Amyloid fibrils are associated with fatal diseases such as Alzheimer's, and amyloid-like fibrils can be formed in vitro. For the yeast protein Sup35, conversion to amyloid-like fibrils is associated with a transmissible infection akin to that caused by mammalian prions. A seven-residue peptide segment from Sup35 forms amyloid-like fibrils and closely related microcrystals, from which we have determined the atomic structure of the cross-beta spine. It is a double beta-sheet, with each sheet formed from parallel segments stacked in register. Side chains protruding from the two sheets form a dry, tightly self-complementing steric zipper, bonding the sheets. Within each sheet, every segment is bound to its two neighbouring segments through stacks of both backbone and side-chain hydrogen bonds. The structure illuminates the stability of amyloid fibrils, their self-seeding characteristic and their tendency to form polymorphic structures.

Amides↗
PubMed2005

Protein crystallography microdiffraction.

Protein microdiffraction using monochromatic beams is becoming a routine tool at third-generation synchrotron radiation sources. Beam sizes have reached the scale of about 5 microm, with illuminated crystal volumes of approximately 500 microm3, as shown for the case of bovine rhodopsin, which was refined to a resolution of 2.6 A. Progress in X-ray optical systems and instrumentation will enable the method to be extended to smaller beams and smaller crystal volumes.

Amyloid↗
PubMed2004

Amyloid fibril formation from sequences of a natural beta-structured fibrous protein, the adenovirus fiber.

Amyloid fibrils are fibrous beta-structures that derive from abnormal folding and assembly of peptides and proteins. Despite a wealth of structural studies on amyloids, the nature of the amyloid structure remains elusive; possible connections to natural, beta-structured fibrous motifs have been suggested. In this work we focus on understanding amyloid structure and formation from sequences of a natural, beta-structured fibrous protein. We show that short peptides (25 to 6 amino acids) corresponding to repetitive sequences from the adenovirus fiber shaft have an intrinsic capacity to form amyloid fibrils as judged by electron microscopy, Congo Red binding, infrared spectroscopy, and x-ray fiber diffraction. In the presence of the globular C-terminal domain of the protein that acts as a trimerization motif, the shaft sequences adopt a triple-stranded, beta-fibrous motif. We discuss the possible structure and arrangement of these sequences within the amyloid fibril, as compared with the one adopted within the native structure. A 6-amino acid peptide, corresponding to the last beta-strand of the shaft, was found to be sufficient to form amyloid fibrils. Structural analysis of these amyloid fibrils suggests that perpendicular stacking of beta-strand repeat units is an underlying common feature of amyloid formation.

Adenoviridae↗
PubMed2004

Structural evolution of regenerated silk fibroin under shear: combined wide- and small-angle x-ray scattering experiments using synchrotron radiation.

The structural evolution of regenerated Bombyx mori silk fibroin during shearing with a Couette cell has been studied in situ by synchrotron radiation small- and wide-angle x-ray scattering techniques. An elongation of fibroin molecules was observed with increasing shear rate, followed by an aggregation phase. The aggregates were found to be amorphous with beta-conformation according to infrared spectroscopy. Scanning x-ray microdiffraction with a 5 microm beam on aggregated material, which had solidified in air, showed silk II reflections and a material with equatorial reflections close to the silk I structure reflections, but with strong differences in reflection intensities. This silk I type material shows up to two low-angle peaks suggesting the presence of water molecules that might be intercalated between hydrogen-bonded sheets.

Fibroins↗
PubMed2003

Recent developments in micro-diffraction on protein crystals.

Protein crystallography micro-diffraction and micro small-angle X-ray scattering at a third-generation synchrotron radiation source undulator beamline both require a compromise on beam size and beam divergence. Micro small-angle X-ray scattering requires in addition an as-close-as-possible angular approach to the direct beam, which is mainly limited by the beam divergence at the sample position. Both techniques have been developed at the ESRF microfocus beamline in a complimentary way. The development of a dedicated microgoniometer in the frame of an EMBL/ESRF collaboration has revealed the potential of micro-diffraction for protein crystallography and is a step in the quest for user friendly instrumentation.

Equipment Design↗
PubMed2003

Influence of CO2 on the micro-structural properties of spider dragline silk: X-ray microdiffraction results.

The mechanical properties of spider dragline fibres are altered by CO(2) exposure under anaesthetizing conditions during the spinning process. In order to relate these macroscopic changes to a microscopic model, the extrusion of dragline silk was studied by synchrotron radiation microdiffraction. A brief exposure of a female Nephila senigalensis spider to CO(2 )results after an incubation time of less than 7 min in the extrusion of a thread (two fibres) swollen with water. The data are interpreted for a model of crystalline beta-sheet domains containing nanofibrils, which reinforce a network of protein chains. The protein network absorbs water, leaving the nanofibrils unaffected. A continuous flow of CO(2) results in a co-extrusion of a dragline thread and an isotropic silk fraction, which probably has a glycine-rich sequence. Long CO(2) exposure reduces the axial alignment of nanofibrils, presumably due to a partial destruction of the amorphous network.

Animals↗
PubMed2003

Control of crystal size and lattice formation by starmaker in otolith biomineralization.

The stone-like otoliths from the ears of teleost fishes are involved in balance and hearing and consist of calcium carbonate crystallites embedded in a protein framework. We report that a previously unknown gene, starmaker, is required in zebrafish for otolith morphogenesis. Reduction of starmaker activity by injection of modified antisense oligonucleotides causes a change in the crystal lattice structure and thus a change in otolith morphology. The expression pattern of starmaker, along with the presence of the protein on the growing otolith, suggest that the expression levels of starmaker control the shape of the otoliths.

Amino Acid Sequence↗
PubMed2003

The mechanical properties of hydrated intermediate filaments: insights from hagfish slime threads.

Intermediate filaments (IFs) impart mechanical integrity to cells, yet IF mechanics are poorly understood. It is assumed that IFs in cells are as stiff as hard alpha-keratin, F-actin, and microtubules, but the high bending flexibility of IFs and the low stiffness of soft alpha-keratins suggest that hydrated IFs may be quite soft. To test this hypothesis, we measured the tensile mechanics of the keratin-like threads from hagfish slime, which are an ideal model for exploring the mechanics of IF bundles and IFs because they consist of tightly packed and aligned IFs. Tensile tests suggest that hydrated IF bundles possess low initial stiffness (E(i) = 6.4 MPa) and remarkable elasticity (up to strains of 0.34), which we attribute to soft elastomeric IF protein terminal domains in series with stiffer coiled coils. The high tensile strength (180 MPa) and toughness (130 MJ/m(3)) of IF bundles support the notion that IFs lend mechanical integrity to cells. Their long-range elasticity suggests that IFs may also allow cells to recover from large deformations. X-ray diffraction and congo-red staining indicate that post-yield deformation leads to an irreversible alpha-->beta conformational transition in IFs, which leads to plastic deformation, and may be used by cells as a mechanosensory cue.

Animals↗
PubMed2002

Comparative architecture of silks, fibrous proteins and their encoding genes in insects and spiders.

The known silk fibroins and fibrous glues are thought to be encoded by members of the same gene family. All silk fibroins sequenced to date contain regions of long-range order (crystalline regions) and/or short-range order (non-crystalline regions). All of the sequenced fibroin silks (Flag or silk from flagelliform gland in spiders; Fhc or heavy chain fibroin silks produced by Lepidoptera larvae) are made up of hierarchically organized, repetitive arrays of amino acids. Fhc fibroin genes are characterized by a similar molecular genetic architecture of two exons and one intron, but the organization and size of these units differs. The Flag, Ser (sericin gene) and BR (Balbiani ring genes; both fibrous proteins) genes are made up of multiple exons and introns. Sequences coding for crystalline and non-crystalline protein domains are integrated in the repetitive regions of Fhc and MA exons, but not in the protein glues Ser1 and BR-1. Genetic 'hot-spots' promote recombination errors in Fhc, MA, and Flag. Codon bias, structural constraint, point mutations, and shortened coding arrays may be alternative means of stabilizing precursor mRNA transcripts. Differential regulation of gene expression and selective splicing of the mRNA transcript may allow rapid adaptation of silk functional properties to different physical environments.

Alternative Splicing↗
PubMedDate not supplied

Fast intracrystalline hydration of beta-chitin revealed by combined microdrop generation and on-line synchrotron radiation microdiffraction.

Water microdrops of about 50 microm in diameter, generated by an ink-jet system, have been used to hydrate fragments of Pogonophora tubes. In situ X-ray microdiffraction with a beam size of 10 microm was used to follow the structural transformations that affected the crystalline beta-chitin part of the specimens. Starting from anhydrous chitin, the formation of a full beta-chitin dihydrate was observed within about 90 s. A disordered intermediate phase with variable d-spacing that could be due to a mixture of anhydrous and hydrated beta-chitin layers was also detected.

Chitin↗
PubMedDate not supplied

Structural study of poly(L-lactic acid) spherulites.

Spherulites of poly(L-lactic acid) (PLLA) and of its blends with atactic poly(3-hydroxybutyrate) (a-PHB, from 10 to 75 wt %) were investigated by microfocus X-ray diffraction using synchrotron radiation. Radial scans in 5 microm steps with 3 microm beam diameter were performed. In PLLA, tens of identical diffraction images were collected. The unit cell a-axis was radially oriented, and the other axes lacked any specific orientation. In contrast, all PLLA/a-PHB blends showed a periodic change of diffraction pattern with increasing distance from the spherulite center. In all cases, the a-axis lay along the radius, while the b- and c-axes rotated about a with a defined periodicity. The unit cell twisting frequency increased with a-PHB content and closely matched the band spacing observed by polarized optical microscopy, which changed from 250 to 60 microm when the amount of a-PHB increased from 10 to 75 wt %. Concomitantly, a gradual broadening of all X-ray reflections was observed.

Crystallization↗