Electron microscopic study of compactization of individual DNA molecules in films during the interaction with histone H1]

The protein-free method was applied for the investigation of histone H1 DNA complexes formation. The main advantage of this method is the possibility to get intramolecular compact structures at interaction of individual spread molecules of DNA with histone H1. It was shown that in the presence of 0.2-5 micrograms/ml of histone H1 in hypophase there are three types of structures on electronmicroscopic preparations: fibres of non-compacted DNA, compact fibres with twisted strands of duplex DNA and compacted rod-like and circular structures where separate fibres of duplex DNA could not be distinguished. The study of compact structures morphology allows to conclude that they are formed by side-by-side association of DNA fibres, as it takes place in the case of triple rings formation at the compactization of circular DNA due to trivaline binding. At increasing ionic strength there is a tendency for transition from second type structures to the third type structures. The latter can be explained by transition from non-cooperative to cooperative binding of histone H1 to DNA.     

Electron microscopic study of changes in chromosomal structural organization under the effect of formamide]

Isolated human metaphase chromosomes were treated with formamide at different (0-70%) concentrations and examined electronmicroscopically by protein monolayer technique. At increasing formamide concentration chromosomes gradually decondense, the scaffold becomes more clearly visible, the loops of chromatin fibres coming off the central part of chromosomes lose their nucleosomal appearance. Electrophoretic analysis of chromosomal proteins data show that formamide-treated chromosomes have approximately the same histone content as those before treatment, although chromosomes treated with 70% formamide look very similar to histone-depleted ones described elsewhere.     

Electron microscopic study of isolated proteoglycans]

Chemically isolated separate preparations of the non-aggregating protein-chondroitin-keratin sulphate (PCKS) fraction from the hyaline cartilage and hyaluronic acid (HUA) of the vitreous body and of the umbilicus were investigated by electron microscopy. PCKS and HUA in films without cytochrome c were present in the form of granules and differed by structural organization. The proteoglycans of the cytochrome c films were seen as finely-filamentous-cellular network, and were distinctly differentiated by their macromolecular organization. A mixture of both proteoglycans formed complexes as a result of a noncovalent interaction. Uranyl acetate ensured a good contrasting of proteoglycans, especially of PCKS, without cytochrome c.     

Electron microscopic study of equine herpesvirus type 1 DNA.

Electron microscopic studies of equine herpesvirus DNA revealed that single strands that were allowed to reanneal formed single-stranded loops with double-stranded stems only at one end of the molecule. These observations support restriction enzyme analyses which indicate that the 92-megadalton DNA molecule exists as a long region of unique sequences covalently linked to a short region. The short region is comprised of an internal unique sequence, which forms the loop during reannealing of single strands, and two terminal inverted repeat sequences that bracket the unique sequence and form the double-stranded stem structure observed upon reannealing of single strands. Measurements of the unique sequence and terminal inverted repeat subgenomic sequences indicate a size of 6.4 megadaltons for each and thus fix the size of the short region at approximately 19.2 megadaltons.     

Oversupercoiling and compactization of supercoiled DNA

Supercoiled DNA pGEMEX with length of 3993 nucleotides was immobilized on the different substrates (freshly cleaved mica, standard aminomica and modified aminomica) and visualized by atomic force microscopy. Plectonomically supercoiled DNA molecules as well as molecules with extremely high level of compactization (i.e. molecules with considerably higher supercoiled density values in comparing with experimentally measured and theoretically investigated ones) were visualized on modified aminomica. At the further increasing of the compactization level an axis length of oversupercoiled molecules was decreased from approximately 390 nm to approximately 140 nm and formation of minitoroids of approximately 50 nm diameter and molecules in sphere conformation were observed. Model of possible conformational transitions of supercoiled DNA was proposed basing on the analysis of captured AFM images at the increasing of supercoiling density.     

Electron microscopic study of DNA complexes with proteins from the Archaebacterium Sulfolobus acidocaldarius

DNA-protein complexes formed in vitro with isolated DNA-binding proteins from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius were analyzed by electron microscopy. Two of the proteins (10a and 10b) formed specific structures after incubation with DNA. Protein 10a, at low protein concentrations, showed individual small spots on the DNA and at high concentrations evenly covered doublestranded DNA. Protein 10b showed three different types of regular structures: one with single-stranded and two with double-stranded DNA. Using double-stranded DNA, 10b first bound cooperatively to two strands forming long, plait-like structures only slightly shorter than respective free DNA. The complex consists of two right-handed, interwound fibers, with a helical pitch of 26 nm and a diameter of ~10-11 nm. At higher protein concentration than needed to package all DNA into the complex with two double-stranded DNAs, the two DNAs were separated again and a new structure was formed evenly covering only one DNA strand. Both structures showed no significant contraction of the length of the DNA covered in the complex. Nucleoprotein formed with single-stranded ΦX174 DNA had a diameter of ~11 nm and could form circles with a contour length of 0.5 μm.     

Electron microscopic study of troponin

Troponin and its components or fragments were observed in an electron microscope by the use of the rotary shadowing technique. In freshly prepared troponin with low viscosity, globular particles were mainly observed. The size of the long axis of the particles was 13.2 +/- 1.3 nm and the size perpendicular to the long axis was 9.5 +/- 1.2 nm. The mean axial ratio was 1.4 +/- 0.3. Most of the particles observed in a stored troponin preparation, having a higher viscosity than that of fresh troponin, had a globular head with a thin tail, with the total length of 25.4 +/- 1.4 nm (head-tail type particles). The axial size of the globular portion was 8.3 +/- 1.2 nm and the tail length was 17.1 +/- 1.6 nm. Observation of various particles during the transitional stages indicated that, in the globular particles, the tail region of head-tail type particle was associated along the globular head region. Troponin T was a filamentous particle with 16.9 +/- 1.5 nm length. The 26K fragment of troponin T, which was devoid of the N-terminal 45 residues from troponin T, was a filamentous particle with the length of 14.4 +/- 1.3 nm. Troponin T1, one of two chymotryptic subfragments of troponin T, was a filamentous particle of 11.6 +/- 1.4 nm length. Troponin C.T in the presence of Ca2+ was a particle with a globular head (7 nm in size) and a tail of about 17 nm length. The Fab fragment of anti-troponin T1 formed regular transverse striations along the thin filament of rabbit skeletal muscle with a 38 nm period.(ABSTRACT TRUNCATED AT 250 WORDS)     

Electron microscopic study of the interaction between cytolytic T-lymphocytes and target cells]

Thymocytes stimulated in vitro in a mixed culture was sorbed by centrifugation on the surface of target cells for electron microscope study of cytology of immune T-lymphocytes and early cytolysis periods. A well developed Golgi apparatus was revealed in the cytoplasm of lymphocytes; there was also accumulation of tubular structures 50 to 60 nm in diameter which communicated with the cysterns of the granular endoplasmic reticulum, with "descended" vesiculi and plasmatic membrane of lymphocyte. This membrane formed numerous contacts with the membrane of target cells thus producing closed clefts. Taking into consideration these data and also current views on the intertransformation of membranes and intracellular transport a hypothetic scheme of the mechanism involved in the cytolysis of the target cell by immune T-lymphocyte was put forward.     

Electron microscopic study of the innervation of the heart conduction system in rats]

A systemic quantitative electron microscopic analysis on innervation of the sinus node, the atrioventricular node, the bundle of His and its pedicles within the interventricular septum has been performed in intact hearts of mature rats. The data have been obtained on the size of nonmyelinated and myelinated nerve fibres, efferent and afferent terminals within different parts of the cardiac conductive system, their interconnection with specialized cardiomyocytes have been described. Application of certain methods for electron microscopic investigation on the innervation of mammalian cardiac conductive system has been discussed.     

Electron microscopic study of the in vitro effect of dipyridamol on the pseudorabies virus

Dipyridamole at a concentration of 50 microM/ml displays no activity on adsorption and penetration of pseudorabies virus in chicken embryonal cells. Furthermore, first stages of virus replication take place within the nucleus, whereas incomplete virus cores defective in DNA content are found within the nucleoplasm at times when the regular viral replication has been finished in controls. Defective pseudorabies virus particles lacking in DNA-content of the core, can be observed at the end of normal replication time. Consequently, the antiviral activity of dipyridamole may be due to blocking of the synthesis or of the incorporation of infectious viral DNA into the virus core.     

Electron microscopic study of alpha-actinin.

Electron microscopic studies of the structure of purified α-actinin alone and in complex with F-actin have determined the molecular shape and size of this protein. α-Actinin molecules represent rods of about 300 Å in length and about 20 Å in diameter.