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H G Wittmann

Publications and source records attributed to H G Wittmann.

At least 37 records · Page 2Linked to original sources
PubMed1987

Three-dimensional crystals of ribosomes and their subunits from eu- and archaebacteria.

Ordered three-dimensional crystals of 70S ribosomes as well as of 30S and 50S ribosomal subunits from various bacteria (E. coli, Bacillus stearothermophilus, Thermus thermophilus and Halobacterium marismortui) have been grown by vapour diffusion in hanging drops using mono- and polyalcohols. A new compact crystal form of 50S subunits has been obtained, and it is suitable for crystallographic studies at medium resolution. In addition, from one crystal form large crystals could be grown in X-ray capillaries. In all cases the crystals were obtained from functionally active ribosomal particles, and the particles from dissolved crystals retained their integrity and biological activity.

Archaea↗
PubMed1987

Reconstitution and crystallisation experiments with isolated split proteins from Bacillus stearothermophilus ribosomes.

Six proteins (B-L1, B-L6, B-L10, B-L11, B-L12 and B-L16) were removed from 50S ribosomal subunits of Bacillus stearothermophilus by treatment with ethanol and ammonium chloride. The proteins were isolated in a pure form, and one of them (B-L6) was crystallized. Five of the six proteins (in various combinations) were added back to the core particles, resulting in 50S subunits lacking one protein. The biological activities of these ribosomal particles as determined in the poly(U)-system varied over a wide range, depending on the protein which was omitted. The particles lacking one protein provide useful tools for heavy-atom derivation necessary for our crystallographic studies on the 50S subunits of Bacillus stearothermophilus.

Bacterial Proteins↗
PubMed1986

The growth of ordered two-dimensional sheets of 70 S ribosomes from Bacillus stearothermophilus.

Well ordered two-dimensional sheets of intact 70 S ribosomes from Bacillus stearothermophilus have been obtained in vitro using salt-alcohol mixtures. These sheets consist of relatively small unit cells with dimensions of 200 +/- 20 A and 400 +/- 30 A. Diffraction patterns of electron micrographs of these sheets stained with uranyl acetate contain features to 42 A resolution.

Geobacillus stearothermophilus↗
PubMed1986

Characterization of single crystals of the large ribosomal particles from a mutant of Bacillus stearothermophilus.

Single, three-dimensional crystals of 50 S ribosomal subunits, from a mutant of Bacillus stearothermophilus that lacks the protein L11, have been characterized using a synchrotron X-ray source. The crystals of the mutated particles grow under the same conditions and are isomorphous to those of the wild type of the same bacteria. They are orthorhombic, contain at least one 2-fold screw axis, and have unit cell dimensions of a = 350(+/- 10) A, b = 670(+/- 10) A, and c = 910(+/- 10) A. They diffract to 15 to 18 A resolution at 4 degrees C and are stable in the synchrotron beam for several hours.

Crystallography↗
PubMed1986

Characterization of single crystals of the large ribosomal particles from Bacillus stearothermophilus.

Single, three-dimensional crystals of the 50 S ribosomal subunit from Bacillus stearothermophilus (strain NCA) have been characterized using a synchrotron X-ray source. The crystals are orthorhombic with unit cell dimensions: a = 350 A, b = 670 A, c = 905 A, and contain at least one 2-fold screw axis. With cooling to -2 degrees C, the large crystals (1.0 mm X 0.2 mm X 0.1 mm) diffract to 15 to 18 A resolution and are stable in the synchrotron beam for several hours. Despite the large cell dimensions, the reflections are readily resolved when the X-ray diffraction patterns are densitometered with a 25 microns faster.

Crystallization↗
PubMed1984

Some x-ray diffraction patterns from single crystals of the large ribosomal subunit from Bacillus stearothermophilus.

X-ray diffraction patterns of three-dimensional crystals of the large ribosomal subunit from Bacillus stearothermophilus have been obtained using a synchrotron radiation beam. The patterns contain resolved diffraction spots and indicate packing in relatively small unit cells, the dimensions of which have been tentatively determined. The internal order of the crystals, as reflected in these patterns, has been correlated with the size, shape and conditions of growth of these crystals.

Geobacillus stearothermophilus↗
PubMed1984

Two-dimensional crystalline sheets of Bacillus stearothermophilus 50S ribosomal particles.

Two-dimensional crystalline sheets of the large ribosomal subunit from Bacillus stearothermophilus have been obtained using a slightly modified procedure to that for growing three-dimensional crystals of the same material. The crystalline subunits are packed within monolayers in a relatively small unit cell, the dimensions of which are closely related to those observed for two forms of the three-dimensional crystals. The packing symmetry is p121, and the optical diffraction patterns of micrographs of negatively stained crystals extend to approximately 3.0 nm.

Crystallization↗
PubMed1984

Ribosomal proteins: their structure and spatial arrangement in prokaryotic ribosomes.

During the last 15 years of ribosomal protein study, enormous progress has been made. Each of the proteins from E. coli ribosomes has been isolated, sequenced, and immunologically and physically characterized. Ribosomal proteins from other sources (e.g., from some bacteria, yeast, and rat) have been isolated and studied as well. Several proteins have recently been crystallized, and from the X-ray studies it is expected that much important information on the three-dimensional structure will be forthcoming. Many other proteins can probably be crystallized if suitable preparative procedures and crystallization conditions are found. Tremendous progress has also been made in deciphering the architecture of the ribosome. A battery of different methods has been used to provide the nearest neighbor distances of the ribosomal proteins in situ. Definitive measurements are now emanating from neutron-scattering experiments which also promise to give reasonably accurate radii of gyration of the proteins in situ. In turn, refined immune electron microscopy results supplement the neutron-scattering data and also position the proteins on the subunits themselves. This cannot be done by the other methods. Determination of the three-dimensional RNA structure within the ribosome is still in its infancy. Nonetheless, it is expected that by combining the data from protein-RNA and from RNA-RNA cross-linking studies, the structure of the RNA in situ can be unraveled. Of great interest is the fact that ribosomal subunits and ribosomes themselves have now been crystallized, and low-resolution structural maps have already been obtained. However, to grow suitable crystals and to resolve the ribosomal structure at a sufficiently high resolution remains a great challenge and task to biochemists and crystallographers.

Amino Acid Sequence↗
PubMed1983

A compact three-dimensional crystal form of the large ribosomal subunit from Bacillus stearothermophilus.

A new form of well-ordered three-dimensional crystals of intact 50 S ribosomal subunits from Bacillus stearothermophilus have been obtained. Electron micrographs of positively stained sections of these crystals revealed that the ribosomal particles are packed closely. The cell parameters have been determined. Representative electron micrographs and their computed contoured filtered images are shown.

Crystallization↗
PubMed1982

Structure and evolution of ribosomes.

Ribosomes are the only cell organelles occurring in all organisms. E. coli ribosomes, which are the best characterized particles, consist of three RNAs and 53 proteins. All components have been isolated and characterized by chemical, physical and immunological methods. The primary structures of the RNAs and of all the proteins are known. Information about the secondary structure of the proteins derives from circular dichroism measurements and from secondary structure prediction methods. The tertiary structure is being studied by limited proteolysis, proton magnetic resonance and crystallization followed by X-ray analysis. Various methods are being used to elucidate the architecture of the ribosomal particle: three-dimensional image reconstruction of crystals of bacterial ribosomes and/or their subunits; immune electron microscopy; neutron scattering; protein-protein, protein-RNA and RNA-RNA crosslinking; total reconstitution of ribosomal subunits. The results from these studies yield valuable information on the architecture of the ribosomal particle. Many mutants have been isolated in which one or a few ribosomal proteins are altered or even deleted. The genetic and biochemical characterization of these mutants allows conclusions about the importance of these proteins for the function of the ribosome. Ribosomal proteins from various prokaryotic and eukaryotic species have been compared by two-dimensional gel electrophoresis, immunological methods, reconstitution and amino acid sequence analysis. These studies show a strong homology among prokaryotic ribosomal proteins but only a weak homology between proteins from prokaryotic and eukaryotic ribosomes. Comparison of the primary and secondary structures of the ribosomal RNAs from various organisms shows that the secondary structure of the RNA molecules has been strongly conserved throughout evolution.

Animals↗
PubMed1982

Parameters for crystal growth of ribosomal subunits.

A systematic analysis of the parameters that control the crystal growth of the large subunit of ribosomes from B stearothermophilus has been carried out. Several parameters have been identified and classified according to their significance. It was found that only biologically active particles can crystallize and that the critical period for the crystallization process is the first few days, during which changes in the volume and content of the crystallization drop are of importance for both nucleation and crystal growth. Consequently, an experimental procedure for fine control of these variables has been developed. As a result of these studies, the reproducibility of crystal formation was increased, and larger and more stable crystals have been obtained.

Bacillus↗
PubMed1981

Macrolide and aminoglycoside antibiotic resistance mutations in the bacillus subtilis ribosome resulting in temperature-sensitive sporulation.

Mutants of Bacillus subtilis resistant to various macrolide antibiotics have been isolated and characterized with respect to their sporulation phenotype and the electrophoretic mobility of their ribosomal proteins (r-proteins). Two types of major alterations of r-protein L17, one probably due to a small deletion, are found among mutants exhibiting high-level macrolide resistance. These mutants are all temperature-sensitive for sporulation (Spots). Low-level resistance to some macrolides is found to be associated with minor alterations in r-protein L17. These mutations do not cause a defective sporulation phenotype. All of the macrolide resistance mutations map at the same locus within the Str-Spc region of the B. subtilis chromosome. Hence, changes in a single ribosomal protein can result in different sporulation phenotypes. Mutants resistant to the aminoglycoside antibiotics neomycin and kanamycin have been isolated. Approximately 5% of these are Spots. Representative mutations, neo162 and kan25, cause concomitant drug resistance and sporulation temperature-sensitivity and map a single-site lesions in the Str-Spc region of the chromosome. Strains bearing neo162 or kan25 are equally cross-resistant to streptomycin or spectinomycin. These mutations define a new B. subtilis drug resistance locus at which mutation can cause defective sporulation.

Aminoglycosides↗