Three dimensional structural studies of α-N-acetylgalactosaminidase (α-NAGA) in α-NAGA deficiency (Kanzaki disease): different gene mutations cause peculiar structural changes in α-NAGAs resulting in different substrate specificities and clinical phenotypes

2005 ◽  
Vol 37 (1) ◽  
pp. 15-20 ◽  
Author(s):  
Takuro Kanekura ◽  
Hitoshi Sakuraba ◽  
Fumiko Matsuzawa ◽  
Seiichi Aikawa ◽  
Hirofumi Doi ◽  
...  
Author(s):  
James A. Lake

The understanding of ribosome structure has advanced considerably in the last several years. Biochemists have characterized the constituent proteins and rRNA's of ribosomes. Complete sequences have been determined for some ribosomal proteins and specific antibodies have been prepared against all E. coli small subunit proteins. In addition, a number of naturally occuring systems of three dimensional ribosome crystals which are suitable for structural studies have been observed in eukaryotes. Although the crystals are, in general, too small for X-ray diffraction, their size is ideal for electron microscopy.


Author(s):  
Jane K. Rosenthal ◽  
Dianne L. Atkins ◽  
William J. Marvin ◽  
Penny A. Krumm

To comprehend structural changes in cardiac myocytes accompanying adrenergic innervation, it is essential that a three dimensional analysis be performed. To date, biological studies which utilize stereological methods have been limited to cells in tissue and in organs. Our laboratory has utilized current stereological techniques for measuring absolute volumes of individual myocytes in primary culture. Cell volumes are calculated for two distinct groups of cells at 96 hours in culture: isolated myocytes and myocytes innervated with adrenergic neurons (Figure 1).Cardiac myocytes are cultured from the ventricular apices of newborn rats. Cells are plated directly onto tissue culture dishes with or without preplated explants from the paravertebral thoracolumbar sympathetic chain. On day four cultures are photographed and marked for one-to-one cell location. Following conventional fixation and embeddment in eponate-12, the cells are relocated and mounted for microtomy. The cells are completely sectioned at 120nm in their parallel orientation to the surface of the dish (Figure 2). Serial sections are collected on formvar coated slotted grids and are recorded in sequence.


Author(s):  
Badrinath Roysam ◽  
Hakan Ancin ◽  
Douglas E. Becker ◽  
Robert W. Mackin ◽  
Matthew M. Chestnut ◽  
...  

This paper summarizes recent advances made by this group in the automated three-dimensional (3-D) image analysis of cytological specimens that are much thicker than the depth of field, and much wider than the field of view of the microscope. The imaging of thick samples is motivated by the need to sample large volumes of tissue rapidly, make more accurate measurements than possible with 2-D sampling, and also to perform analysis in a manner that preserves the relative locations and 3-D structures of the cells. The motivation to study specimens much wider than the field of view arises when measurements and insights at the tissue, rather than the cell level are needed.The term “analysis” indicates a activities ranging from cell counting, neuron tracing, cell morphometry, measurement of tracers, through characterization of large populations of cells with regard to higher-level tissue organization by detecting patterns such as 3-D spatial clustering, the presence of subpopulations, and their relationships to each other. Of even more interest are changes in these parameters as a function of development, and as a reaction to external stimuli. There is a widespread need to measure structural changes in tissue caused by toxins, physiologic states, biochemicals, aging, development, and electrochemical or physical stimuli. These agents could affect the number of cells per unit volume of tissue, cell volume and shape, and cause structural changes in individual cells, inter-connections, or subtle changes in higher-level tissue architecture. It is important to process large intact volumes of tissue to achieve adequate sampling and sensitivity to subtle changes. It is desirable to perform such studies rapidly, with utmost automation, and at minimal cost. Automated 3-D image analysis methods offer unique advantages and opportunities, without making simplifying assumptions of tissue uniformity, unlike random sampling methods such as stereology.12 Although stereological methods are known to be statistically unbiased, they may not be statistically efficient. Another disadvantage of sampling methods is the lack of full visual confirmation - an attractive feature of image analysis based methods.


Author(s):  
Gregory J. Czarnota

Chromatin structure at the fundamental level of the nucleosome is important in vital cellular processes. Recent biochemical and genetic analyses show that nucleosome structure and structural changes are very active participants in gene expression, facilitating or inhibiting transcription and reflecting the physiological state of the cell. Structural states and transitions for this macromolecular complex, composed of DNA wound about a heterotypic octamer of variously modified histone proteins, have been measured by physico-chemical techniques and by enzyme-accessibility and are recognized to occur with various post-translational modifications, gene activation, transformation and with ionic-environment. In spite of studies which indicate various forms of nucleosome structure, all current x-ray and neutron diffraction studies have consistently resulted in only one structure, suggestive of a static conformation. In contrast, two-dimensional electron microscopy studies and three-dimensional reconstruction techniques have yielded different structures. These fundamental differences between EM and other ultrastructural studies have created a long standing quandary, which I have addressed and resolved using spectroscopic electron microscopy and statistical analyses of nucleosome images in a study of nucleosome structure with ionic environment.


Author(s):  
B.V.V. Prasad ◽  
E. Marietta ◽  
J.W. Burns ◽  
M.K. Estes ◽  
W. Chiu

Rotaviruses are spherical, double-shelled particles. They have been identified as a major cause of infantile gastroenteritis worldwide. In our earlier studies we determined the three-dimensional structures of double-and single-shelled simian rotavirus embedded in vitreous ice using electron cryomicroscopy and image processing techniques to a resolution of 40Å. A distinctive feature of the rotavirus structure is the presence of 132 large channels spanning across both the shells at all 5- and 6-coordinated positions of a T=13ℓ icosahedral lattice. The outer shell has 60 spikes emanating from its relatively smooth surface. The inner shell, in contrast, exhibits a bristly surface made of 260 morphological units at all local and strict 3-fold axes (Fig.l).The outer shell of rotavirus is made up of two proteins, VP4 and VP7. VP7, a glycoprotein and a neutralization antigen, is the major component. VP4 has been implicated in several important functions such as cell penetration, hemagglutination, neutralization and virulence. From our earlier studies we had proposed that the spikes correspond to VP4 and the rest of the surface is composed of VP7. Our recent structural studies, using the same techniques, with monoclonal antibodies specific to VP4 have established that surface spikes are made up of VP4.


1999 ◽  
Vol 82 (08) ◽  
pp. 277-282 ◽  
Author(s):  
Yuri Veklich ◽  
Jean-Philippe Collet ◽  
Charles Francis ◽  
John W. Weisel

IntroductionMuch is known about the fibrinolytic system that converts fibrin-bound plasminogen to the active protease, plasmin, using plasminogen activators, such as tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator. Plasmin then cleaves fibrin at specific sites and generates soluble fragments, many of which have been characterized, providing the basis for a molecular model of the polypeptide chain degradation.1-3 Soluble degradation products of fibrin have also been characterized by transmission electron microscopy, yielding a model for their structure.4 Moreover, high resolution, three-dimensional structures of certain fibrinogen fragments has provided a wealth of information that may be useful in understanding how various proteins bind to fibrin and the overall process of fibrinolysis (Doolittle, this volume).5,6 Both the rate of fibrinolysis and the structures of soluble derivatives are determined in part by the fibrin network structure itself. Furthermore, the activation of plasminogen by t-PA is accelerated by the conversion of fibrinogen to fibrin, and this reaction is also affected by the structure of the fibrin. For example, clots made of thin fibers have a decreased rate of conversion of plasminogen to plasmin by t-PA, and they generally are lysed more slowly than clots composed of thick fibers.7-9 Under other conditions, however, clots made of thin fibers may be lysed more rapidly.10 In addition, fibrin clots composed of abnormally thin fibers formed from certain dysfibrinogens display decreased plasminogen binding and a lower rate of fibrinolysis.11-13 Therefore, our increasing knowledge of various dysfibrinogenemias will aid our understanding of mechanisms of fibrinolysis (Matsuda, this volume).14,15 To account for these diverse observations and more fully understand the molecular basis of fibrinolysis, more knowledge of the physical changes in the fibrin matrix that precede solubilization is required. In this report, we summarize recent experiments utilizing transmission and scanning electron microscopy and confocal light microscopy to provide information about the structural changes occurring in polymerized fibrin during fibrinolysis. Many of the results of these experiments were unexpected and suggest some aspects of potential molecular mechanisms of fibrinolysis, which will also be described here.


2018 ◽  
Vol 1 (1) ◽  
pp. 1-11 ◽  
Author(s):  
Kamaljit Singh Boparai ◽  
Rupinder Singh

This study highlights the thermal characterization of ABS-Graphene blended three dimensional (3D) printed functional prototypes by fused deposition modeling (FDM) process. These functional prototypes have some applications as electro-chemical energy storage devices (EESD). Initially, the suitability of ABS-Graphene composite material for FDM applications has been examined by melt flow index (MFI) test. After establishing MFI, the feedstock filament for FDM has been prepared by an extrusion process. The fabricated filament has been used for printing 3D functional prototypes for printing of in-house EESD. The differential scanning calorimeter (DSC) analysis was conducted to understand the effect on glass transition temperature with the inclusion of Graphene (Gr) particles. It has been observed that the reinforced Gr particles act as a thermal reservoir (sink) and enhances its thermal/electrical conductivity. Also, FT-IR spectra realized the structural changes with the inclusion of Gr in ABS matrix. The results are supported by scanning electron microscopy (SEM) based micrographs for understanding the morphological changes.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rui Zhai ◽  
Hui Chen ◽  
Zhihua Shan

AbstractElectrochemical modification of animal skin is a new material preparation method and new direction of research exploration. In this study, under the action of the electric field using NaCl as the supporting electrolyte, the effect of electrolysis on Glycyl-glycine(GlyGl), gelatin(Gel) and Three-dimensional rawhide collagen(3DC) were determined. The amino group of GlyGl is quickly eliminated within the anode region by electrolysis isolated by an anion exchange membrane. Using the same method, it was found that the molecular weight of Gel and the isoelectric point of the Gel decreased, and the viscosity and transparency of the Gel solution obviously changed. The electrolytic dissolution and structural changes of 3DC were further investigated. The results of TOC and TN showed that the organic matter in 3DC was dissolved by electrolysis, and the tissue cavitation was obvious. A new approach for the preparation of collagen-based multi-pore biomaterials by electrochemical method was explored.


2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Angika Bulbul ◽  
Joseph Rosen

AbstractPartial aperture imaging system (PAIS) is a recently developed concept in which the traditional disc-shaped aperture is replaced by an aperture with a much smaller area and yet its imaging capabilities are comparable to the full aperture systems. Recently PAIS was demonstrated as an indirect incoherent digital three-dimensional imaging technique. Later it was successfully implemented in the study of the synthetic marginal aperture with revolving telescopes (SMART) to provide superresolution with subaperture area that was less than one percent of the area of the full synthetic disc-shaped aperture. In the study of SMART, the concept of PAIS was tested by placing eight coded phase reflectors along the boundary of the full synthetic aperture. In the current study, various improvements of PAIS are tested and its performance is compared with the other equivalent systems. Among the structural changes, we test ring-shaped eight coded phase subapertures with the same area as of the previous circular subapertures, distributed along the boundary of the full disc-shaped aperture. Another change in the current system is the use of coded phase mask with a point response of a sparse dot pattern. The third change is in the reconstruction process in which a nonlinear correlation with optimal parameters is implemented. With the improved image quality, the modified-PAIS can save weight and cost of imaging devices in general and of space telescopes in particular. Experimental results with reflective objects show that the concept of coded aperture extends the limits of classical imaging.


2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Michael Zenzes ◽  
Paul Zaslansky

AbstractMicro-CT provides critical data for musculoskeletal research, yielding three-dimensional datasets containing distributions of mineral density. Using high-resolution scans, we quantified changes in the fine architecture of bone in the spine of young mice. This data is made available as a reference to physiological cancellous bone growth. The scans (n = 19) depict the extensive structural changes typical for female C57BL/6 mice pups, aged 1-, 3-, 7-, 10- and 14-days post-partum, as they attain the mature geometry. We reveal the micro-morphology down to individual trabeculae in the spine that follow phases of mineral-tissue rearrangement in the growing lumbar vertebra on a micrometer length scale. Phantom data is provided to facilitate mineral density calibration. Conventional histomorphometry matched with our micro-CT data on selected samples confirms the validity and accuracy of our 3D scans. The data may thus serve as a reference for modeling normal bone growth and can be used to benchmark other experiments assessing the effects of biomaterials, tissue growth, healing, and regeneration.


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