Structural Homology between Prokaryotic and Eukaryotic Ribosomes

1977 ◽  
Vol 35 ◽  
pp. 416-417
Author(s):  
M. Boublik ◽  
W. Hellmann ◽  
F. Jenkins
Keyword(s):  
Ribosomal Proteins ◽  
Critical Evaluation ◽  
Three Dimensional ◽  
Structural Complexity ◽  
Carbon Support ◽  
Uranyl Acetate ◽  
Three Dimensional Model ◽  
Ribonucleic Acids ◽  
E Coli ◽  
70S Ribosome

The structural complexity of ribosomes is evident from the large number of their constituents - ribosomal proteins and ribonucleic acids. The simplest known prokaryotic (70S) ribosome from E. coli is composed of 53 different proteins and three ribonucleic acid molecules (23S, 16S and 5S RNA). Eukaryotic (80S) ribosomes also consist of three RNAs (28S, 18S and 5S) but contain a higher number of proteins, approximately 70. The mutual interactions of ribosomal proteins and RNAs determine the final structure and thus the biological activity of the ribosome. As expected for particles with the above composition, ribosomes and both ribosomal subunits have no symmetry or repetition in structure.Electron micrographs of ribosomes (stained with 0.5% aqueous uranyl acetate and deposited on a fine carbon support) show distinct contours and electron dense pattern. Critical evaluation of different views of the two-dimensional projections of the ribosomal contours can provide a reasonable basis for proposing a three-dimensional model of the ribosome.

Structural similarity of ribosomes from E. coli and A. salina

1978 ◽  
Vol 36 (2) ◽  
pp. 242-243
Author(s):  
M. Boublik ◽  
R.M. Wydro ◽  
W. Hellmann ◽  
F. Jenkins
Keyword(s):  
Ribosomal Proteins ◽  
Direct Evidence ◽  
Structural Similarity ◽  
Carbon Support ◽  
Artemia Salina ◽  
Uranyl Acetate ◽  
Biological Assays ◽  
E Coli ◽  
And Function ◽  
Fine Carbon

Ribosomes are ribonucleoprotein particles necessary for processing the genetic information of mRNA into proteins. Analogy in composition and function of ribosomes from diverse species, established by biochemical and biological assays, implies their structural similarity. Direct evidence obtained by electron microscopy seems to be of increasing relevance in understanding the structure of ribosomes and the mechanism of their role in protein synthesis.The extent of the structural homology between prokaryotic and eukaryotic ribosomes has been studied on ribosomes of Escherichia coli (E.c.) and Artemia salina (A.s.). Despite the established differences in size and in the amount and proportion of ribosomal proteins and RNAs both types of ribosomes show an overall similarity. The monosomes (stained with 0.5% aqueous uranyl acetate and deposited on a fine carbon support) appear in the electron micrographs as round particles with a diameter of approximately 225Å for the 70S E.c. (Fig. 1) and 260Å for the 80S A.s. monosome (Fig. 2).

Ribosome Structural Organization

1974 ◽  
Vol 32 ◽  
pp. 332-333
Author(s):  
James A. Lake
Keyword(s):  
Ribosomal Proteins ◽  
Structural Organization ◽  
Three Dimensional ◽  
Small Subunit ◽  
Structural Studies ◽  
X Ray Diffraction ◽  
Specific Antibodies ◽  
X Ray ◽  
E Coli ◽  
Complete Sequences

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.

Structural Role of rRNAs on the Ribosomal Subunits from E. Coli by Scanning Transmission Electron Microscopy

1981 ◽  
Vol 39 ◽  
pp. 424-425
Author(s):  
M. Boublik ◽  
N. Robakis ◽  
J.S. Wall
Keyword(s):  
Three Dimensional ◽  
Uranyl Acetate ◽  
Dimensional Structure ◽  
Electron Microscopic ◽  
Ribosomal Subunits ◽  
E Coli ◽  
Freeze Dried ◽  
16S Rna ◽  
Scanning Transmission ◽  
And Function

The three-dimensional structure and function of biological supramolecular complexes are, in general, determined and stabilized by conformation and interactions of their macromolecular components. In the case of ribosomes, it has been suggested that one of the functions of ribosomal RNAs is to act as a scaffold maintaining the shape of the ribosomal subunits. In order to investigate this question, we have conducted a comparative TEM and STEM study of the structure of the small 30S subunit of E. coli and its 16S RNA.The conventional electron microscopic imaging of nucleic acids is performed by spreading them in the presence of protein or detergent; the particles are contrasted by electron dense solution (uranyl acetate) or by shadowing with metal (tungsten). By using the STEM on freeze-dried specimens we have avoided the shearing forces of the spreading, and minimized both the collapse of rRNA due to air drying and the loss of resolution due to staining or shadowing. Figure 1, is a conventional (TEM) electron micrograph of 30S E. coli subunits contrasted with uranyl acetate.

scholarly journals Structure of the Bacterial Ribosome at 2 Å Resolution

2020 ◽  
Author(s):  
Zoe L. Watson ◽  
Fred R. Ward ◽  
Raphaël Méheust ◽  
Omer Ad ◽  
Alanna Schepartz ◽  
...  
Keyword(s):  
Ribosomal Proteins ◽  
Noncovalent Interactions ◽  
Ribosomal Subunit ◽  
E Coli ◽  
Bacterial Ribosome ◽  
Domains Of Life ◽  
Chemical Resolution ◽  
70S Ribosome ◽  
A Site ◽  
First Time

AbstractContinuing advances in cryo-electron microscopy (cryo-EM) demonstrate the promise it holds for revealing biological structures at chemical resolution, in which noncovalent interactions, RNA and protein modifications, and solvation can be modeled accurately. At present, the best cryo-EM-derived models of the bacterial ribosome are of the large (50S) ribosomal subunit with effective global resolutions of 2.4-2.5 Å, based on map-to-model Fourier shell correlation (FSC). Here we present a model of the E. coli 70S ribosome with an effective global resolution of 2.0 Å, based on maps showcasing unambiguous positioning of residues, their detailed chemical interactions, and chemical modifications. These modifications include the first examples of isopeptide and thioamide backbone substitutions in ribosomal proteins, the former of which is likely conserved in all domains of life. The model also defines extensive solvation of the small (30S) ribosomal subunit for the first time, as well as interactions with A-site and P-site tRNAs, mRNA, and the antibiotic paromomycin. The high quality of the maps now allows a deeper phylogenetic analysis of ribosomal components, and identification of structural conservation to the level of solvation. The maps and models of the bacterial ribosome presented here should enable future structural analysis of the chemical basis for translation, and the development of robust tools for cryo-EM structure modeling and refinement.

Morphology of escherichia coli ribosomes as obtained by high resolution shadow cast

1978 ◽  
Vol 36 (2) ◽  
pp. 240-241 ◽  
Author(s):  
B. Tesche ◽  
G.W. Tischendorf ◽  
G. Stöffler
Keyword(s):  
Freeze Drying ◽  
Three Dimensional ◽  
Optical Resolution ◽  
Single Particles ◽  
Three Dimensional Model ◽  
Rna Molecules ◽  
E Coli ◽  
Fine Grained ◽  
Shadow Casting ◽  
Shadow Cast

All components (54 proteins and 3 RNA molecules) of the E. coli ribosome have been identified and characterized by biochemical and biophysical means. The morphology of the ribosome and the structural arrangement of its components, however, is not yet unequivocally known. The reasons for this are that crystal arrangements could not be obtained and that the different preparations of single particles remain invariant; and that holds for different laboratories as well.We applied a preparation which has been demonstrated to be most gentle and hence most structure-preserving; that is, freeze-drying with shadow casting at -150°C. Furthermore, the evaporator is of such design that it produces very little heat and renders a thin, fine-grained tungsten layer. An electron optical resolution of 6 Å can be achieved. Fig. 1 shows a micrograph of a typical shadowgraph obtained with 70S ribosomes from which we attempt to derive a three-dimensional model of the ribosome.

scholarly journals Mutagenesis in the α3α4 GyrA Helix and in the Toprim Domain of GyrB Refines the Contribution of Mycobacterium tuberculosis DNA Gyrase to Intrinsic Resistance to Quinolones

2008 ◽  
Vol 52 (8) ◽  
pp. 2909-2914 ◽  
Author(s):  
Stéphanie Matrat ◽  
Alexandra Aubry ◽  
Claudine Mayer ◽  
Vincent Jarlier ◽  
Emmanuelle Cambau
Keyword(s):  
Escherichia Coli ◽  
Mycobacterium Tuberculosis ◽  
Primary Structure ◽  
Three Dimensional ◽  
Dna Gyrase ◽  
Quinolone Resistance ◽  
Dimensional Model ◽  
Three Dimensional Model ◽  
Intrinsic Resistance ◽  
E Coli

ABSTRACT The replacement of M74 in GyrA, A83 in GyrA, and R447 in GyrB of Mycobacterium tuberculosis gyrase by their Escherichia coli homologs resulted in active enzymes as quinolone susceptible as the E. coli gyrase. This demonstrates that the primary structure of gyrase determines intrinsic quinolone resistance and was supported by a three-dimensional model of N-terminal GyrA.

scholarly journals Three-Dimensional Model for the Human Cl−/HCO3− Exchanger, AE1, by Homology to the E. coli ClC Protein

2013 ◽  
Vol 425 (14) ◽  
pp. 2591-2608 ◽  
Author(s):  
Pamela Bonar ◽  
Hans-Peter Schneider ◽  
Holger M. Becker ◽  
Joachim W. Deitmer ◽  
Joseph R. Casey
Keyword(s):  
Three Dimensional ◽  
Dimensional Model ◽  
Three Dimensional Model ◽  
E Coli

scholarly journals Enzymatic and structural properties of human glutamine:fructose-6-phosphate amidotransferase 2 (hGFAT2)

2020 ◽  
pp. jbc.RA120.015189
Author(s):  
Isadora A. Oliveira ◽  
Diego Allonso ◽  
Tácio V. A. Fernandes ◽  
Daniela M.S. Lucena ◽  
Gustavo T. Ventura ◽  
...  
Keyword(s):  
Biosynthetic Pathway ◽  
Three Dimensional ◽  
Dimensional Model ◽  
Expression And Purification ◽  
Three Dimensional Model ◽  
Hexosamine Biosynthetic Pathway ◽  
E Coli ◽  
Cellular Processes ◽  
Catalytically Active ◽  
Dynamics Simulations

Glycoconjugates play a central role in several cellular processes and alteration in their composition is associated with numerous human pathologies. Substrates for cellular glycosylation are synthesized in the hexosamine biosynthetic pathway, which is controlled by the glutamine:fructose-6-phosphate amidotransfera-se (GFAT). Human isoform 2 GFAT (hGFAT2) has been implicated in diabetes and cancer; however, there is no information about structural and enzymatic properties of this enzyme. Here, we report a successful expression and purification of a catalytically active recombinant hGFAT2 (rhGFAT2) in E. coli cells fused or not to a HisTag at the C-terminal end. Our enzyme kinetics data suggest that hGFAT2 does not follow the expected ordered bi-bi mechanism, and performs the glucosamine-6-phosphate synthesis much more slowly than previously reported for other GFATs. In addition, hGFAT2 is able to isomerize fructose-6-phosphate into glucose-6-phosphate even in the presence of equimolar amounts of glutamine, which results in unproductive glutamine hydrolysis. Structural analysis of a three-dimensional model of rhGFAT2, corroborated by circular dichroism data, indicated the presence of a partially structured loop in the glutaminase domain, whose sequence is present in eukaryotic enzymes but absent in the E. coli homolog. Molecular dynamics simulations suggest that this loop is the most flexible portion of the protein, and plays a key role on conformational states of hGFAT2. Thus, our study provides the first comprehensive set of data on the structure, kinetics and mechanics of hGFAT2, which will certainly contribute to further studies on the (patho)physiology of hGFAT2.

scholarly journals Updates on enzymatic and structural properties of human glutamine: fructose-6-phosphate amidotransferase 2 (hGFAT2)

2020 ◽  
Author(s):  
Isadora A. Oliveira ◽  
Diego Allonso ◽  
Tácio V. A. Fernandes ◽  
Daniela M. S. Lucena ◽  
Gustavo T. Ventura ◽  
...  
Keyword(s):  
Biosynthetic Pathway ◽  
Three Dimensional ◽  
Dimensional Model ◽  
Expression And Purification ◽  
Three Dimensional Model ◽  
Hexosamine Biosynthetic Pathway ◽  
E Coli ◽  
Cellular Processes ◽  
Catalytically Active ◽  
Dynamics Simulations

AbstractGlycoconjugates play a central role in several cellular processes and alteration in their composition is associated to human pathologies. The hexosamine biosynthetic pathway is a route through which cells obtain substrates for cellular glycosylation, and is controlled by the glutamine: fructose-6-phosphate amidotransferase (GFAT). Human isoform 2 GFAT (hGFAT2) has been implicated in diabetes and cancer, however, there is no information about structural and enzymatic properties of this enzyme. Here, we report a successful expression and purification of a catalytically active recombinant hGFAT2 (rhGFAT2) in E. coli cells fused or not to a HisTag at the C-terminal end. Our enzyme kinetics data suggest that hGFAT2 does not follow the ordered bi-bi mechanism, and performs the glucosamine-6-phosphate synthesis much slowly than previously reported for other GFATs. In addition, hGFAT2 is able to isomerase fructose-6-phosphate into glucose-6-phosphate even in presence of equimolar amounts of glutamine, in an unproductive glutamine hydrolysis. Structural analysis of the generated three-dimensional model rhGFAT2, corroborated by circular dichroism data, indicated the presence of a partially structured loop in glutaminase domain, whose sequence is present in eukaryotic enzymes but absent in the E. coli homolog. Molecular dynamics simulations show such loop as the most flexible portion of the protein, which interacts with the protein mainly through the interdomain region, and plays a key role on conformational states of hGFAT2. Altogether, our study provides the first comprehensive set of data on the structure, kinetics and mechanics of hGFAT2, which will certainly contribute for further studies focusing on drug development targeting hGFAT2.

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