Regular Aggregates Research Articles

Trivaline molecular complexes with trinucleotides form in solution the extended, about 1000 A in length structures.

The fluorescence, flow linear dichroism and electron microscopy (EM) have shown the trivaline ability to interact in solution with certain molecules of trinucleotides. This interaction results in formation of extended structures up to several thousand angstroms in length. Such structures were observed for trivaline complexes with homopurine, homopyrimidine or random sequences of deoxyribo- and ribonucleotides, independently of the presence or absence of the terminal 5'-phosphate residue. A model of such a structural organization is proposed. An elementary structural unit consists of a trivaline beta-dimer and adsorbed trinucleotide. So, "dimeric" complex is formed. Two such "dimeric" complexes combine with each other by means of peptide-peptide contacts (as with beta-sandwich). So, "tetrameric" complex is formed. It has a dyad axis. Two such structural units combine with each other by means of Hoogsteen's hydrogen bonds. So, "octameric" complex is formed. It has three mutually perpendicular dyad axes. The "octameric" complexes appear to be able to combine with each other by means of stacking interactions, and to form the regular organized aggregates consisting of many dozens of elementary units. So, "stacking" structure is formed. The "octameric" complex is the symmetry translational unit of such a structure. The spatial position of the bases in all these structures is additionally fixed by the nucleo-peptide interactions. These aggregates have the appearance of extended structures on electron micrographs.

A polypeptide of 59 kDa is associated with bundles of cytoplasmic filaments in Neurospora crassa

Complex arrangements of filamentous structures have been isolated from vegetative cells of the fungus Neurospora crassa. They were enriched by differential centrifugation and purified by permeation chromatography. The filamentous structures are made up of units of 8-10 nm diameter and were isolated in bundles of up to six to nine units. The main constituent of these structures is a polypeptide with an apparent molecular mass of 59 kDa (P59Nc), which represents 4-5% of the total N. crassa proteins. The filamentous structures are cold-stable and are not affected by high-ionic-strength solutions or by the presence of 10 mM-EDTA or 1% (w/v) Triton X-100; they were disassembled by raising the pH of the solution or by using Tris-based buffers. The disassembled form assembled into structures sedimentable at 105,000 g after dialysis against the isolation buffer. The sedimentable structures were organized in the form of regular aggregates of 42-45 nm polypeptides and reacted weakly with anti-IFA, a monoclonal antibody which recognizes an epitope common to many of the higher-eukaryote intermediate-filament polypeptides. Immunofluorescence examination of wall-digested hyphae of N. crassa using affinity-purified antibodies prepared against P59Nc showed immunostaining of abundant filamentous and dot-shaped structures distributed in the cytoplasm.

Expression of tartrate-resistant acid phosphatase in B-CLL treated with phorbol ester or phorbol ester plus calcium ionophore.

Cells from 3 patients with B-chronic lymphocytic leukaemia (B-CLL) and 1 with B-prolymphocytic leukaemia (B-PLL) were treated in vitro with the phorbol ester 12-0-tetradecanoylphorbol (TPA), the calcium ionophore A23187, or a combination of TPA and A23187. TPA induced the cells to adhere to the culture flask or to clump in dense clusters; single cells became enlarged, often with cytoplasmic elongations. Cells treated with TPA plus A23187 acquired a plasmacytoid morphology and formed regular aggregates in culture. Only TPA alone induced the expression of tartrate-resistant acid phosphatase (TRAP) as documented by isoelectric focusing on horizontal thin-layer gels. The TRAP isoenzyme was first detected after 24 h of TPA treatment; its intensity increased during further TPA exposure, being maximally expressed at 72/96 h. The results suggest that, while TPA triggers B-CLL cells to convert to hairy cell leukaemia (HCL)-type cells, the double stimulus which more closely imitates physiological activation initiates a 'normal' differentiation programme which leads to plasma cells.

Structure of helical RecA-DNA complexes: II. Local conformational changes visualized in bundles of RecA-ATPγS filaments

Complexes of RecA-DNA filaments, formed in the presence of a non-hydrolyzable ATP analog, ATPγS, aggregate together into regular bundles in the presence of Mg 2+. Electron micrographs of several different forms of RecA-double-stranded DNA bundles have been analyzed: bundles of six supercoiled filaments at two different concentrations of Mg 2+, and bundles of three supercoiled filaments at a single concentration of Mg 2+. The bundles are all characterized by a regular left-handed supercoiling of the component filaments arising from the non-integral number of RecA subunits per turn of the RecA helix in these aggregates, about 6.15 units/turn. When single-stranded DNA is used instead of double-stranded DNA, regular aggregates composed of many filaments are formed. These aggregates do not supercoil, consistent with a symmetry of the component filaments of close to 6.0 units/turn. These different structures have provided a strong confirmation of the analysis of isolated RecA filaments. Since different RecA protomers within the component filaments of these aggregates are in different environments, they have provided a direct view of different conformations that RecA subunits may adopt within the same filament as a result of non-equivalent contacts. The conformational changes we have visualized are quite large, with apparent movements of mass over distances greater than 2 nm. The RecA-mediated strand exchange reaction is a highly dynamic process, which involves both the unwinding and stretching of DNA, in addition to the physical movement of DNA strands. It is quite likely, therefore, that the different conformations of RecA subunits seen in these aggregates represent different states of RecA during its enzymatic strand exchange activity.

Structural and functional evidence for multiple channel complexes in the outer membrane of Neurospora crassa mitochondria.

The outer membrane of mitochondria contains proteins that form channels called VDAC (voltage-dependent anion-selective channels). Two independent lines of evidence suggest that these channels occur in specific complexes in the outer membrane of Neurospora mitochondria. Electron microscopic images of these outer membranes reveal polymorphic crystalline arrays of putative pores. These arrays can be shown to be interrelated by movement in the membrane plane of a particular rigid channel triplet or of regular aggregates of this triplet. Detergent extracts of the same outer membranes induce single- and multiple-step conductances in planar phospholipid membranes, with a marked preference for the insertion of triplets and multiples of triplets. The tendency of the mitochondrial channels to occur as extended arrays, apparently built up from triplets, may have important functional and evolutionary implications.

The variation of intermolecular contacts in helical aggregates of glutamine synthetase

Glutamine synthetase from E. coli is a large (MW 600,000) oligomeric enzyme composed of twelve identical polypeptide chains of MW 50,000. These twelve subunits are arranged at the vertices of two eclipsed hexagons to produce a structure with symmetry 622 or D6 as shown by electron microscopy and X-ray diffraction. The dodecameric molecule has a hexagonal profile of approximately 140 Å diameter and a thickness of approximately 90 Å. When treated with divalent cations such as Co++, Zn++, Ni++ or Cu++, glutamine synthetase molecules aggregate along their six-fold axes to form long strands and these strands will often wrap around one another to form cables (Fig. 1). The aggregation is completely reversed by the addition of excess Mn++ resulting in regeneration of full enzymatic activjjy. The most regular helical aggregates are formed in the presence of 10 mM Co++ at pH 7.0, and we have used these helical cables to study the three dimensional structure and symmetry of the glutamine synthetase molecule and the types of protein-protein interactions involved in the formation of glutamine synthetase cables.

A review of statistical structures in synthetic polypeptides and biological macromolecules

The relationship between the structure and function of biological macromolecules is fundamental to our understanding of molecular biology. It is generally true that those proteins which are rich in charged residues have a high axial ratio (fibrous proteins) and those which are low in charged residues tend to have a small axial ratio (globular proteins). Rod-shaped proteins have a larger surface area and smaller internal volume per unit volume than do the globular proteins. The surface charge per unit area is reduced by the protein adopting a cylindrical shape. Also, a larger percentage of apolar residues can be accommodated in the centre of a globular unit than in a cylindrical unit. This review is primarily concerned with attempting to understand how the long, rod-shaped molecules found in fibrous proteins pack together in three dimensions in a regular or quasi-regular manner to produce functionally useful filaments or fibrils such as are present in muscle, tendon, cartilage, skin etc. Since it is often possible to produce synthetic filaments by self-assembly of the long molecules from solution, the structure of the molecular aggregate must be specifically related to the packing of the individual molecules. Electron microscopy has allowed the visualization of orderly arrays of filaments in muscle and hair but technical limitations still prevent easy recognition of regular packing of the molecules themselves. X-ray diffraction, however, conclusively shows the existence of such regular assemblies of molecules in the native state of many biological fibres. Long rodshaped macromolecules are specifically designed by nature to aggregate. The kind of molecular interactions in the aggregate are normally dictated by thermodynamic considerations of free energy. In the event of the possible formation of several structures of similar free energy, that which is biologically useful must be preferred. The purpose of a structural investigation of a fibrous protein is to see how the molecular structure can lead to specific interactions which produce regular aggregates and also how these interactions bestow properties appropriate to the biological role on the resulting aggregate. In this review we draw attention to a particular aspect of regular arrays of fibrous proteins, their synthetic analogues and also the nucleic acids, viz. statisticality. We first describe the various possible categories of statisticality and show how the X-ray diffraction patterns from such structures may be calculated. Then follows a review of the known occurrences of statisticality in assemblies of biopolymers and their analogues and a discussion of the biological implications. We also point out the characteristic features of X-ray diffraction patterns from structures which exhibit statistical ordering.

A study of Chromobacterium typhiflavum from the rat.

SummaryTwo strains of an organism possessing unusual morphological features have been isolated from the rat. Aggregations of individual cells into discrete spherical ‘large bodies’ are found. These bodies appear in chain forms. The nature of the matrix binding the cells is not known. Individual young cells are motile by peritrichous flagella, swarm in soft agar and are Gram negative. Sections of cells show a typically Gram negative cell wall and mode of division. The cell surface appears convoluted in the Gram negative manner. Well defined buds continuous with the cell wall have been demonstrated and many cells contain crystalloid and peripheral bodies. The organisms ferment a wide variety of carbohydrates and are aerobic and facultative anaerobes. These organisms appear to fit the description of Chromobacterium typhiflavum; however the ability to form regular aggregates, containing cells distinct from the ‘vegetative’ form, and to form buds would indicate that this organism should be placed in a new genus.

Geometry of regular aggregates

Geometry of regular aggregates

Origin of rigid bacterial "giant flagella" ("giant whips", riesenzöpfe).

Development of "giant flagella" in microcultures of Proteus vulgaris occurred by an aggregation of flagella from motionless cells in a microcolony. Regular free flagellar aggregations very similar to those of the Proteus also were encountered in microcultures of a peritrichous Bacillus species, but were seen in but one of a very large number of microcultures of a monotrichous Escherichia coli strain. Observation of actively motile cells in wet mounts and of stained preparations of the three species with dark phase contrast, Nomarski differential interference, dark-field, and brightfield optics led to an hypothesis concerning the life history which accepts the flagellum as being flexible in the functional state. After an active period extending through more than one generation, the flagellum becomes rigid and apparently is ejected from the cell as a complete unit, including what appears to be a bulbous basal body.

The systemic infection of an aphid by a plant virus

Electron microscopy of aphids ( Hyperomyzus lactucae L.) that had acquired lettuce necrotic yellows virus (LNYV) from infected sow thistle ( Sonchus oleraceus L.) revealed two types of bacilliform particles. One type appeared identical to those reported from plant cells and from purified LNYV preparations, and the other was similar but lacked an outer coat. No particles were seen in aphids reared on virus-free plants or in embryos developing within infective aphids. In the mature aphid the characteristic particles were found in the muscles, fat body, brain, mycetome, tracheae, epidermis, salivary glands and in the cells of the alimentary canal. The large number of particles forming regular aggregates in some of the cells of these organs is interpreted as evidence for virus multiplication within the aphid.

The forced magnetostriction of single crystal gadolinium

Complexes of RecA-DNA filaments, formed in the presence of a non-hydrolyzable ATP analog, ATPγS, aggregate together into regular bundles in the presence of Mg2+. Electron micrographs of several different forms of RecA-double-stranded DNA bundles have been analyzed: bundles of six supercoiled filaments at two different concentrations of Mg2+, and bundles of three supercoiled filaments at a single concentration of Mg2+. The bundles are all characterized by a regular left-handed supercoiling of the component filaments arising from the non-integral number of RecA subunits per turn of the RecA helix in these aggregates, about 6.15 units/turn. When single-stranded DNA is used instead of double-stranded DNA, regular aggregates composed of many filaments are formed. These aggregates do not supercoil, consistent with a symmetry of the component filaments of close to 6.0 units/turn. These different structures have provided a strong confirmation of the analysis of isolated RecA filaments. Since different RecA protomers within the component filaments of these aggregates are in different environments, they have provided a direct view of different conformations that RecA subunits may adopt within the same filament as a result of non-equivalent contacts. The conformational changes we have visualized are quite large, with apparent movements of mass over distances greater than 2 nm. The RecA-mediated strand exchange reaction is a highly dynamic process, which involves both the unwinding and stretching of DNA, in addition to the physical movement of DNA strands. It is quite likely, therefore, that the different conformations of RecA subunits seen in these aggregates represent different states of RecA during its enzymatic strand exchange activity.

THE STRUCTURE OF AGGREGATES AND THE MOLECULAR KINEMATICS OF THE VISCOSITY OF A BERNAL LIQUID

The structure of 3-dimensional aggregates is discussed as a set of points on which graphs are constructed. By constructing the Voronoi honeycomb (Dirichlet regions) for the points and applying a small "irregularizing transformation", a "simplicial graph" and a "primitive coordination number" (whose value is close to 14 for all aggregates) can be defined universally for both regular and irregular aggregates. Recent studies of the geometry of irregular aggregates (of steel balls, crystal grains, etc.) are reviewed. The theory of liquids of J. D. Bernal is discussed and the simplicial graph is used to show that the "activation volume" of a Bernal liquid is about one-tenth of the molecular volume. The kinematics of flow of aggregates is discussed in terms of their graphs and in terms of a process of "volume exchange"—the production and destruction of free volume. Using these concepts, an equation is derived for the viscosity of a Bernal liquid as a product of five terms expressing respectively the kinematic, stoichiometric, kinetic, pressure-dependent, and shear-dependent factors.

Crystallinity of Calcium Chondroitin Sulphate

WELL-DEFINED macroscopic shapes containing symmetry elements are generally regarded as a sign of crystalline arrangement of molecules. Recently, Kargin1 demonstrated that polymer molecules might have a regular geometrical aggregate as viewed in the electron microscope without being crystalline so far as X-ray diffraction measurement is concerned and vice versa. In our work with calcium chondroitin sulphate A we encountered apparently a similar phenomenon. Einbinder and Schubert2 reported that calcium chondroitin sulphate (their so-called mid and tail fraction) showed crystalline shapes under the microscope when precipitated with ethanol from aqueous solution. These same fractions, however, proved to be amorphous in our X-ray diffraction experiments. Further investigation also revealed that the ‘head’ fraction of calcium chondroitin sulphate which Einbinder and Schubert2 reported as amorphous under the microscope was highly crystalline in X-ray diffraction studies. Detailed analysis of these results will be reported elsewhere.

A theory of devitrification of igneous rocks

That the devitrification of igneous rock is brought about by regular aggregations or arrangements of the molecules of the substance, is evident from the fact that a vitreous rock which has the same composition as a holocrystalline variety always exhibits a lower specific gravity than the latter. For instance, if we compare equal bulks of obsidian and granite, two rocks which have practically identical qualitative and quantitive chemical compositions, we find that the former is the lighter of the two, both, of course, as regards weight in air and weight in water. This solitary fact is all that is necessary to prove that in the glassy obsidian there is less matter, and consequently a smaller number of molecules present than in an equal volume of its holocrystalline relative. If this be so, then the difference in the number of those infinitesimally minute units in each case must be due to the manner in which they are respectively arranged. From the study of crystal development we know that the arrangement of molecules in crystalline matter must occur on some rigidly regular principle, and that it is to this regularity of arrangement that the form, optical properties, cleavage, and other important physical characters of crystals are wholly due. The plasma or mother liquor out of which the crystals grow, must, on the other hand, have its constituent molecules scattered about in some purely higgledy-piggledy fashion, as it exhibits properties the very opposite of those of crystalline material, and is, moreover, bulk for