Study of human erythrocyte membrane proteins by 2-dimensional electrophoresis

Fractionation of human erythrocyte membrane proteins was performed using a modification of two-dimensional gel electrophoresis described by P. O'Farrel with isoelectric point plotted against molecular mass. All major erythrocyte proteins, including high molecular weight proteins, such as spectrin and band 3 protein, identified by one-dimensional sodium dodecyl sulfate gel electrophoresis, were visualized by silver staining of two-dimensional gels. All in all about 50 polypeptides were distinguished on two-dimensional electrophoretic patterns. Preliminary protein map was developed.     

Changes in the ionization equilibrium of erythrocyte suspension under heating]

The effects of ionic strength, urea, calcium and fluorine ions, ouabain and cholinesterase inhibitors on the changes in the ionization equilibrium of an erythrocyte suspension under heating were studied. Proton release by erythrocytes was compared to a release of potassium ions and hemoglobin from the cells. The proton release under heating is mainly determined by the physico--chemical properties of superficial structures of erythrocytes and does not depend on the activity of cholinesterase, ATPase and glycolytic processes.     

Effect of modifiers of natural biological response on the functional activity of macrophages (oncological aspect)

Induction of cytotoxicity by biological response modifiers (BRMs) is only one aspect of macrophage activation. After the use of BRMs there were other changes in the functional activity of cells and in particular their increased production or secretion of a number of growth factors. Thus, activation of macrophage antitumor activity induced by BCG vaccine was transitory while activation of growth factor production was more stable in time which finally led to increased proliferation of tumor cells. Combined use of cyclophosphamide and BCG vaccine significantly increased not only the toxicity induced by BCG vaccine but also their liberation of the growth factors. Such macrophages lost their ability to control the growth of a small number of the tumor cells cultivated in their presence. Development of ways for directed activation of macrophages aimed at elimination of the tumor cells which survived the chemotherapy should include evaluation of the combined effect of various BRMs and chemotherapeutics on both antitumor and protumor activity i. e. ability to produce the factors stimulating the tumor growth.     

Evolutionary relationships of eukaryotic kingdoms

The evolutionary relationships of four eukaryotic kingdoms—Animalia, Plantae, Fungi, and Protista—remain unclear. In particular, statistical support for the closeness of animals to fungi rather than to plants is lacking, and a preferred branching order of these and other eukaryotic lineages is still controversial even though molecular sequences from diverse eukaryotic taxa have been analyzed. We report a statistical analysis of 214 sequences of nuclear small-subunit ribosomal RNA (srRNA) gene undertaken to clarify these evolutionary relationships. We have considered the variability of substitution rates and the nonindependence of nucleotide substitution across sites in the srRNA gene in testing alternative hypotheses regarding the branching patterns of eukaryote phylogeny. We find that the rates of evolution among sites in the srRNA sequences vary substantially and are approximately gamma distributed with size and shape parameter equal to 0.76. Our results suggest that (1) the animals and true fungi are indeed closer to each other than to any other crown group in the eukaryote tree, (2) red algae are the closest relatives of animals, true fungi, and green plants, and (3) the heterokonts and alveolates probably evolved prior to the divergence of red algae and animal-fungus-green-plant lineages. Furthermore, our analyses indicate that the branching order of the eukaryotic lineages that diverged prior to the evolution of alveolates may be generally difficult to resolve with the srRNA sequence data.     

A survey of toxicity of cyanobacterial blooms in Lake Ladoga and adjacent water bodies

Twentyfive cyanobacterial blooms in Lake Ladoga and adjacent water bodies were studied in the summer of 1990–1992. Toxicity of the water bloom material for mice was detected in 9 cases. The maximal tolerable doses (MTD) of the material extracted from biomass varied within 3–30 mg kg^–1 mouse body weight; 50% lethal doses (LD₅₀) were within 45–125 mg kg^–1. Toxic water blooms were registered in Karelian lakes and in the Neva Bay, Gulf of Finland. Cyanobacterial samples collected on the eastern coast of Lake Ladoga proved to be non-toxic. The species identified in toxic bloom material included Anabaena circinalis, A. flos-aquae, A. lemmermannii, Anabaena sp., Aphanizomenonflos-aquae, Gloeotrichia echinulata, G. pisum, Microcystis aeruginosa and Oscillatoria sp. These data suggest that toxic forms of cyanobacteria are widespread in Karelian lakes belonging to the drainage basin of Lake Ladoga.     

The role ofSaccharomyces cerevisiae Cdc40p in DNA replication and mitotic spindle formation and/or maintenance

Successful progression through the cell cycle requires the coupling of mitotic spindle formation to DNA replication. In this report we present evidence suggesting that, inSaccharomyces cerevisiae, theCDC40 gene product is required to regulate both DNA replication and mitotic spindle formation. The deduced amino acid sequence ofCDC40 (455 amino acids) contains four copies of a β-transducin-like repeat. Cdc40p is essential only at elevated temperatures, as a complete deletion or a truncated protein (deletion of the C-terminal 217 amino acids in thecdc40-1 allele) results in normal vegetative growth at 23°C, and cell cycle arrest at 36°C. In the mitotic cell cycle Cdc40p is apparently required for at least two steps: (1) for entry into S phase (neither DNA synthesis, nor mitotic spindle formation occurs at 36°C and (2) for completion of S-phase (cdc40::LEU2 cells cannot complete the cell cycle when returned to the permissive temperature in the presence of hydroxyurea). The role of Cdc40p as a regulatory protein linking DNA synthesis, spindle assembly/maintenance, and maturation promoting factor (MPF) activity is discussed.     

The influence of the cytosolic oncotic pressure on the permeability of the mitochondrial outer membrane for ADP: implications for the kinetic properties of mitochondrial creatine kinase and for ADP channelling into the intermembrane space

Summary Cytosolic proteins as components of the physiological mitochondrial environment were substituted by dextrans added to media normally used for incubation of isolated mitochondria. Under these conditions the volume of the intermembrane space decreases and the contact sites between the both mitochondrial membranes increase drastically. These morphological changes are accompanied by a reduced permeability of the mitochondrial outer compartment for adenine nucleotides as it was shown by extensive kinetic studies of mitochondrial enzymes (oxidative phosphorylation, mi-creatine kinase, mi-adenylate kinase). The decreased permeability of the mitochondrial outer membrane causes increased rate dependent concentration gradients in the micromolar range for adenine nucleotides between the intermembrane space and the extramitochondrial space. Although all metabolites crossing the outer membrane exhibit the same concentration gradients, considerable compartmentations are detectable for ADP only due to its low extramitochondrial concentration. The consequences of ADP-compartmentation in the mitochondrial intermembrane space for ADP-channelling into the mitochondria are discussed.