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PubMed1977

[Quaternary structure of oligomeric immunoglobulin A forms from human blood serum].

A character of forces stabilyzing quaternary structure of dimer and more high molecular human immunoglobulin A oligomers is found to be different. Quaternary structure of IgA dimer is formed when joining subunits with disulfide bonds and is stabilized by non-covalent interactions between them. Disulfide bonds play a main part in the formation of trimers and tetramers. Dimer IgA reconstructs by 40% from subunits with intact interchain S--S bonds. The addition of exogenous J-chain does not significantly affect the process of dimer self-assembling from subunits with recovered and intact interchain disulfide bonds.

Chemical Phenomena↗
PubMed1979

The binding of ribosomal protein S4 does not change the gross conformation of the 16 S RNA.

The binding of ribosomal protein S4 to the 16 S RNA does not result in a large shape or conformational change in the 16 S RNA under the conditions of reconstitution. The sedimentation coefficient, frictional coefficient ratio, and effective hydrodynamic radius of the 16 S RNA.protein S4 complex are very similar to those obtained for the 16 S RNA free in solution. Only subtle conformational differences were obtained in the comparison of the complex and free 16 S RNA by circular dichroism. Thus, extensive organization of the 16 S RNA by ribosomal protein S4 is not a step in the process of self-assembly of the 30 S subunit.

Circular Dichroism↗
PubMedDate not supplied

[Spectrum differentiation as a method of studying protein preparations with a high turbidity level].

The studies on a model of a mixture of fibrin-monomer with glycogen and of fibrin gels of different turbidity show that in the first and second derivatives of the absorption spectra the contribution of turbidity is several orders lower. The derivative spectra are shown to be valuable technique to study the phenomena that are accompanied by substantial changes in turbidity: protein association and self-assembly of supermolecular structure.

Fibrin↗
PubMed1984

The living skin-equivalent as a structural and immunological model in skin grafting.

The living skin equivalent is a tissue formed when self-assembled collagen fibrils are contracted by fibroblasts. The rate and extent of volume contraction is proportional to the number of cells incorporated into the lattice. The fibroblasts are biochemically active in the lattice, synthesizing collagen and adding it to the matrix. This compacted lattice provides a substrate allowing attachment of keratinocytes and the formation of a multilayered keratinized epidermis with a basement lamella. When a skin equivalent grafted to a recipient animal is wounded, it is capable of wound contraction and repair. Because of the simplified character of the skin equivalent it may offer a useful model for studying wound healing. During the initial healing of the grafted skin equivalent, fibroblasts from the skin equivalent move freely into the surrounding tissue. Karyotyping of fibroblasts grown from skin surrounding the skin equivalent graft shows that fibroblasts from the graft migrate at least 6-9 mm from the graft edge. At 3-6 mm from the graft edge about 50% of the fibroblasts present are of donor origin. Fibroblasts also migrate from allogenic skin grafts before these grafts are rejected. At 3-6 mm from the scar left by the rejected graft 36% of the fibroblasts are of donor origin 2 months after grafting. The skin equivalent model provides a well-defined system for studying transplantation reactions. Results obtained on persistence of allografted fibroblasts support our theory that in the rat, the fibroblast is antigenically neutral.

Animals↗
PubMed1982

[Generation of the differences of the electric potentials by Rhodospirillum rubrum reaction center complexes devoid of the heavy subunit].

The electrogenic activity of Rhodospirillum rubrum P870 reaction center complexes devoid of the heavy (H) subunit and retaining the light (L) and medium (M) subunits, was studied. The proteoliposomes containing such reaction center complexes were formed by a self-assembly procedure, using soya bean phospholipids. In the presence of Ca2+ the reaction center proteoliposomes were incorporated into a phospholipid-impregnated Teflon filter separating two solutions of identical composition. After addition of N,N,N',N'-tetra-methyl-p-phenylenediamine (or cytochrome c) and qinone (menadione), illumination caused generation of an electric potential difference between the two filter-separated compartments, the proteoliposome-free compartment being negatively charged. The illuminated proteoliposomes took up penetrating tetraphenylphosphonium cations and, in a lesser degree, tetraphenylborate anions. The data obtained suggest that the reaction center complexes containing only L- and M-subunits possess the electrogenic activity. The H-subunit is not directly involved in membrane potential generation.

Bacterial Proteins↗
PubMed1984

The mechanism of action of Na glutamate, lysine HCl, and piperazine-N,N'-bis(2-ethanesulfonic acid) in the stabilization of tubulin and microtubule formation.

Preferential interaction measurements between proteins and monosodium glutamate were carried out to arrive at an understanding of the mechanism of its strong effect on tubulin stability and self-assembly into microtubules. For all proteins studied, i.e. bovine serum albumin, lysozyme, beta-lactoglobulin, and calf brain tubulin, the protein showed a large preferential hydration in the presence of monosodium glutamate. The enhancement of tubulin self-association by monosodium glutamate can be interpreted in terms of the large unfavorable free energy of interaction between the additive and the protein. Preferential interactions were also examined for lysine hydrochloride, which also gave a preferential hydration of the proteins, except for tubulin. The dependence of the preferential hydration parameter on proteins was different for the two additives, suggesting the importance of net electrostatic charges of proteins in their interaction with glutamate anions and lysinium cations. The zero preferential interaction of lysine hydrochloride with tubulin indicates an affinity of the lysine cation for the protein. Both additives increased the transition temperature of proteins. This can be understood in terms of the unfavorable free energy of interaction between the additive and the protein surface, which should be even more unfavorable when the denaturation causes an increase in the surface area.

Alkanesulfonates↗
PubMed1984

Mineralization by matrix vesicles.

Matrix vesicles are widely regarded as the initial site of calcification in epiphyseal growth plate cartilage, in growing bone and in predentin. This opinion has recently been challenged on grounds that the early aqueous methods used for electron microscopic tissue preparation may have produced an erroneous picture by causing mineral dislocation. However, this argument has now been refuted by multiple investigators throughout the world using a variety of anhydrous methods coupled with electron probe analysis to show convincingly that matrix vesicles are, indeed, associated with initial mineral. Matrix vesicles appear to mineralize by concentrating calcium and phosphate at a protected site close to the inner leaflet of the vesicle membrane. Calcium may be attracted by its affinity for acidic phospholipids of the vesicle membrane, and phosphate may be concentrated by the action of transmembrane phosphatases of the matrix vesicle membrane. Evidence is accumulating to suggest that alkaline phosphatase of the matrix vesicle membrane functions as a phosphotransferase or phosphate vector, transporting PO4 across the vesicle membrane. The mechanism(s) of matrix vesicle biogenesis are discussed including budding from the plasma membrane (for which there is much support), cell degeneration (for which there is gathering support), extrusion of intracytoplasmic vesicles (for which there is weak support), and extracellular subunit self-assembly (for which there is little support). It is suggested that none of these mechanisms is necessarily exclusive, thus more than one mechanism may function in the same tissue. Finally, it is noted that in many calcific diseases, ranging from arthritis to atherosclerosis, mineralization is initiated by extracellular membrane-invested vesicles which are probably analogous to the matrix vesicles of skeletal tissues.

Animals↗
PubMed1984

[Changes in the ratio of microsomal electron carriers in reconstituted microsomal membranes].

The reconstitution of microsomal membrane monooxygenase system with variable contents of the hydroxylating chain enzymatic components was carried out. It was found that during self-assembly of microsomal membranes solubilized with 4% sodium cholate and gel filtration through Sephadex LH-20 in the presence of isolated microsomal enzymes, two forms of cytochrome P-450, i. e. phenobarbital- and 3-methylcholantrene-induced ones, and NADPH-cytochrome P-450 reductase, the exogenous enzymes are incorporated into the microsomal membrane matrices of control and methyl-cholantrene-treated animals. In the membranes reconstituted from the microsomes of the methylcholantrene-induced animals the catalytic activity of cytochrome P-448 in the metabolism of benz(a)pyrene at varying cytochrome P-448 and NADPH-cytochrome P-450 reductase contents were investigated.

Animals↗
PubMed1984

[Catalytic activity of cytochrome P-448 in native and reconstituted microsomal membranes].

The dependence of oxidative activity of a microsomal monooxygenase system on the ratio of its constituent enzymes--cytochrome P-448 and NADPH-cytochrome P-450 reductase--was investigated. Two approaches for preparation of microsomal membranes with different contents of cytochrome P-448 were used, i. e., time-dependent induction by 3-methylcholanthrene and reconstitution of microsomal membranes by self-assembly from a mixture of solubilized microsomes of intact and methylcholanthrene-induced rats. Using antibodies against cytochrome P-448, the cytochrome P-448 content in native and reconstituted microsomal membranes and its participation in hydroxylation of benz(a)pyrene was determined. It was shown that benz(a)pyrene is hydroxylated in a membrane microsomal monooxygenase system more effectively, when the ratio of NADPH-cytochrome P-450 reductase to cytochrome P-448 is 1: 3 to 1: 6.

Animals↗
PubMed1984

Cholesteric organization of DNA in vivo and in vitro.

In concentrated solutions DNA organizes spontaneously to form the "cholesteric" phase which is one type of liquid crystal. We have reproducibly obtained both continuous cholesteric phases and isolated cholesteric globules in equilibrium with the isotropic phase. A comparison is made between this in vitro cholesteric organization and dinoflagellate chromosomes which present the same organization in vivo. The observed defects are analyzed in the two cases. It appears that the cholesteric organization is due to self-assembly phenomena and that the shape of globules and chromosomes is due both to surface tensions and to the presence of defects.

Chromosomes↗
PubMed1982

Construction of bacterial flagellar filaments, and aspects of their conversion to different helical forms.

The helical flagellar filaments of bacteria such as Salmonella typhimurium are constructed from subunits of a single protein called flagellin by a process of self-assembly. They are polymorphic, and change from one helical form to another in a variety of circumstances. This paper discusses, from a viewpoint of classical mechanics, two types of problem associated with these filaments. First is the general problem of how to construct a helical rod from identical subunits. There must be some variation in the pattern of packing of subunits; and it is demonstrated that a mechanically bi-stable but otherwise linear-elastic subunit can build filaments which are not only helical but also possess the observed polymorphic properties. The question of uniqueness of the design is discussed. The second problem concerns the hydrodynamic performance of a helical filament which can switch waveform when subjected to mechanical overload. Transitions which can be brought about in this way are reviewed briefly. An explanation is offered in terms of the mechanics of polymorphism for the behaviour of an anomalous and puzzling 'partly rotating' filament in Hotani's (1979) micro-video study of tethered filaments in fluid streams.

Bacteria↗
PubMed1980

[Structural organization of membranes reconstituted from phospholipids and subchromatophore pigment-protein complexes].

Pigment--protein complexes of the P870 reaction centers and complexes of the bacteriochlorophyll light-harvesting antenna were isolated from the chromatophores of the non-sulfur purple bacterium Rhodospirillum rubrum by solubilization with detergents. The proteoliposomes containing the reaction centers or reaction centers and the light-harvesting antenna as well as liposomes formed from phospholipids were obtained by a self-assembly procedure using seya bean phospholipids. The freeze-fracture study showed that the proteoliposomes contain a large amount of globular particles. The particles incorporated into the two types of the proteoliposomes were distinguished in size. The globules of the reaction center and antenna complexes were bigger in size than the reaction center globules. The globular structures were not found in the liposomal membranes. The liposomes formed in the absence of the pigment--protein complexes were predominantly the multilamellar vesicles. The proteoliposomes were mostly represented as monolamellar membrane vesicles. The spatial arrangement of the reaction center complexes in the membranes is discussed.

Bacterial Chromatophores↗
PubMed1983

[Reconstitution of membranous microsomal monooxygenase system during gel filtration on Sephadex LH-20].

Solubilization of liver microsomes of intact, phenobarbital-treated and 3-methylcholantrene-treated rats was investigated. It was shown that treatment of microsomes with 4% sodium cholate leads to separation of proteins and phospholipids from the microsomal membrane. The microsomal solubilizate was passed through lipophilic Sephadex LH-20 and the particles whose properties are close to those of the microsomal ones were obtained. The high efficiency of self-assembly of the microsome-like particles, e. g. lack of cytochrome P-450 conversion into cytochrome P-420, high microsomal protein yield and the high level of metabolizing activity of microsomal monooxygenase, was demonstrated.

Animals↗
PubMed1983

Assembly of unfertilized sea urchin egg tubulin at physiological temperatures.

Tubulin was purified from unfertilized eggs of the sea urchin Strongylocentrotus purpuratus by DEAE-column chromatography and cycles of temperature-dependent assembly and disassembly. Tubulin-containing column fractions self-assemble into intact microtubules in the absence of high molecular weight microtubule-associated proteins. Egg microtubules assembled during the third cycle of assembly following DEAE-chromatography are composed of 2 or 3 alpha tubulins and 2 beta tubulins as assayed by isoelectric focusing and two-dimensional electrophoresis. The critical protein concentrations necessary for assembly of egg tubulin at 37 and 25 degrees C are 0.15-0.24 and 0.24-0.28 mg/ml, respectively. At physiological temperatures, the critical protein concentrations are 0.81 mg/ml at 15 degrees C and 0.70-0.79 mg/ml at 18 degrees C. At 18 degrees C, bovine brain microtubule-associated proteins stoichiometrically stimulate the initial rate and final extent of egg tubulin assembly. These hybrid microtubules assemble at 18 degrees C at a critical protein concentration of 4-20 micrograms/ml.

Animals↗
PubMed1978

Monoclonal IgG anticoagulants delaying fibrin aggregation in two patients with systemic lupus erythematosus (SLE).

There is paucity of information regarding the prolonged plasma thrombin time known to occur in some patients with systemic lupus erythematosus. Detailed investigations of plasma from two such patients disclosed that IgG accounted for this defect in each case. IgG isolated from plasma of either patient possessed the property of delaying fibrin aggregation and prolonging the clotting times of fibrinogen. Preincubation of IgG from either patient with anti-IgG or anti-Fab (rabbit) serum abolished this anticoagulant property. Moreover, the anticoagulant IgG from the first patient was neutralized with anit-k chain and anti-IgG3, that from the second patient with anti-lambda chain and anti-IgG1 serum. These anticoagulants were also dissimilar with respect to their interactions with fibrin(ogen). IgG from the first patient had no anticoagulant activity against fibrin(ogen) species lacking intact Aalpha chains. IgG from the second patient displayed undiminished anticoagulant effect on such fibrin(ogen) species. We conclude that each anticoagulant interacted with a distinct region(s) on the fibrinogen molecule and that these interactions affect or involve sites that participate in the fibrin self-assembly process.

Adolescent↗
PubMed1976

Biogenesis of matrix vesicles in cartilage growth plates.

Although the structure and function of cartilage matrix vesicles have been comprehensively documented in the literature, the method of vesicle production and export to the extra-cellular matrix is less well understood. The existing data are grouped into four general hypotheses of matrix vesicle biogenesis, with a consideration of the relative merits and weaknesses of each postulate. The hypotheses are: a) budding from cellular processes; b) extrusion of preformed structures; c) cellular degeneration and disintegration; and c)subunit secretion and extracellular assembly. Each of these possibilities offers conceptual advantages and disadvantages, so that it is difficult to adopt a monistic stance. Previous notions of vesicle biogenesis have derived from statis morphologic and analytical data, which are necessarily limiting when attempting to draw kinetic conclusions. Radiotracer studies of lipid synthesis and transport in cartilage lead to the conclusion that matrix vesicles in the growth plate are derived from chondrocytes; the present data favor the hypotheses of budding from cellular processes and/or subunit secretion with extracellular self-assembly.

Cartilage↗
PubMed1977

Myogenesis and contraction in the early embryonic heart of the rainbow trout. An electron microscopic study.

Myogenesis in the embryonic heart of the rainbow trout, Salmo gairdneri (Rich.), was investigated electron microscopically from the 29th to the 41st somite stage. Thick and thin myofilaments are formed simultaneously as well as precursors of Z-lines, to which the thin filaments are attached. The genesis of filaments takes place in the region around the intracellular yolk droplets. The first myofibrils appear by the 33rd somite stage, probably formed by a mechanism of self-assembly in which the binding sites of actin and myosin participate. A- and I-bands do not develop before the 38th osmite stage. The contraction already begins during the 33rd somite stage in the middle of the tubular heart. Gradually, the peristaltic waves spread increasingly to other parts of the heart. In the 41st somite stage the entire heart is contractile and all myocytes contain myofibrils.

Animals↗
PubMedDate not supplied

[Determination of fibrin and fibrinogen split products by their anticoagulant activity].

A new diagnostic method to determine fibrinogen and fibrin splitting products is based on the fact that high-molecular fragments acting as specific inhibitors of fibrin self-assembly compose these products. Fibrinogen and fibrin splitting products are detected and determined quantitatively by prolongation of coagulation time in the test with the standard monomeric fibrin. The number of units of anticoagulating activity in the studied urine are the results of the determinations. Sensitivity of the method if necessary may be increased by adding of a certain amount of the fibrinogen and fibrin splitting products to the test that makes a favourable background for the action of the products. The method is rather sensitive, reliable and simple. It may be used in ordinary clinical laboratories. The method proved to be quite satisfactory when testing renal diseases at the clinic. It is of great significance for early detection of the transplanted kidney rejection. The new method is compared with the known immunological method which consist in determination of the inhibitory effect of the fibrinogen and fibrin splitting products on agglutination of erythrocytes of specific antibodies; the erythrocytes being subjected to "tanning" and "loading" with fibrinogen. A satisfactory correlation is found between the results of two methods.

Adult↗