Isolation and Characterization of Humin-Like Substances Produced by Wood-Degrading White Rot Fungi

Three samples of high-molecular-weight humin-like substances were obtained by solid-phase cultivation of Coriolus hirsutus and/or Cerrena maxima on oat straw. The yield of humin-like substances amounted to 1.38–2.26% of the weight of the plant substrate consumed. These substances, produced both by individual and mixed cultures of the basidiomycetes, were shown to be similar in their structure and physicochemical properties. According to the data of IR and ¹³C-NMR spectroscopy, the substances contained aromatic fragments and were close to soil humic acids. Studies of the dynamics of laccase production suggested that the humin-like substances were produced via direct degradation of lignin macromolecules with direct involvement of extracellular laccase.     

Chemotactic effect of urokinase plasminogen activator: a major role for mechanisms independent of its proteolytic or growth factor domains.

Urokinase type plasminogen activator (uPA) converts plasminogen to plasmin and is highly chemotactic for many cell types. We examined, using recombinant wild type and mutated forms of uPA, the extent to which its proteolytic properties, its growth-like domain (GFD) and/or interactions with the specific receptor (uPAR) contribute to the chemotactic activity towards vascular smooth muscle cells (SMC). Recombinant wild type uPA (r-uPA) stimulated cell migration nearly 5.8-fold, inactive r-uPA, with a mutation in the catalitic domain (r-uPA(H/Q)), 3-fold, uPA without growth factor like domain (r-uPA(GFD )), 2.6-fold, and a form containing both mutations (r-uPA(H/Q, GFD ), 3.3-fold. All recombinant forms of uPA, wild type and those with mutations were equally and highly effective (IC50 approximately 20 nM) in displacing 125I-r-uPA bound to SMC. These results indicate that additional mechanisms, not dependent on uPA's proteolytic activity or the binding ability of its GFD to uPAR, are the major contributors to its chemotactic action on SMC.     

Temperature-induced changes in copper centers and protein conformation of two fungal laccases from Coriolus hirsutus and Coriolus zonatus.

The paper reports on two fungal laccases from Coriolus hirsutus and Coriolus zonatus and their type-2 copper-depleted derivatives. Temperature-induced changes of the copper centers were characterized by optical and electron paramagnetic resonance (EPR) spectroscopy, and the overall protein stability by differential scanning microcalorimetry. The intact enzymes showed highly cooperative thermal unfolding transitions at about 90 degrees C. Type-2 copper depletion led to uncoupling of the domains characterized by a different melting pattern which resolved three subtransitions. Melting curves monitored optically at 290, 340 and 610 nm showed additional transitions below thermal unfolding temperature. EPR spectra of the intact laccases showed the disintegration of the trinuclear copper cluster accompanied by loss of one of the copper ions and disappearance of the strong antiferromagnetic coupling in the type-3 site at 70 degrees C and above 70 degrees C. The copper centers of type-2 copper-depleted laccase showed reduced thermotolerance.     

Fluorescence cytochemistry of nucleic acids. The current status and outlook: on the 35th anniversary of the method

Most important fields of the fluorescence cytochemistry of nucleic acids are reviewed, special attention being paid to the following aspects of the problem: physicochemical basis of different methods of staining; new methods of cell analysis, including flow cytometry; functional studies, i.e. the use of fluorescence approaches to obtain information about the functional state of individual cells and cell aggregates.     

The development and structure of the optic nerve in normal embryos and fetuses of the white rat and after x-ray irradiation]

With the aim to reveal the ionising radiation effect to formation and structure of the optic nerve (ON), 55 white rat intact embryos and fetuses and 77 experimental embryos and fetuses, subjected to x-ray irradiation on the 10th-14th day of development, have been studied. The main regularities in formation of the ON have been stated under normal conditions. Certain disturbances in formation of the ON and in the internal membrane of the optic vesicle (future retina) under effect of x-rays in the dose of 2.24 Gy have been detected during the intrauterine period of development--folds, rosellas of retina, retardation in differentiation of the retina nervous layer, aberration of the ON growth with a subsequent reduction, the ON hypo- and aplasia, retardation in formation of neural sheaths, absence of intraspace between the sheaths.     

Erythropoiesis and osteogenesis (II. The mechanism of the suppression of erythropoiesis during the reparative process in bone tissue)

The experiment on rats and mice has shown that the presence of a developing callus in the organism after fracture of the tubular bones inhibits erythropoiesis. Inhibition of erythropoiesis is due to delay of erythropoietin biosynthesis by serotonin. A problem on two-fold serotonin effect is under discussion: on the one hand, serotonin inhibits the erythropoietin formation and delays erythropoiesis, on the other hand, it intensifies the effect of ready erythropoietin on the developing bone marrow erythroid cells.     

Genetic study of plasmid integration into yeast chromosomes. IV. Integration of the plasmid pYF91 into different yeast chromosomes

Integration of the episomic chimeric plasmid pYF91 into yeast chromosomes has been studied. Plasmid insertion into the chromosomes was observed to occur with the frequency of 4 X 10(-8). 379 integrants were selected from the highly unstable (cir0) transformants. The fact of plasmid integration into particular chromosomes was confirmed for 318 integrants. Genetic analysis showed that the plasmid can integrate into the region of LEU2 gene or into another arm of chromosome III (227 integrants), and also into other chromosomes: I, II, IV, V, VI, VII, VIII, IX, XII, XV (91 integrants). It is suggested that integration is the result of recombination between yeast chromosomes and homologous plasmid regions carrying LEU2 gene or Ty element and "delta" sequence.     

Mechanism of the radioprotecting action of chemical compounds on Escherichia coli cells.

Summary The effect of radioprotection of indolylakylamines (5-methoxytryptamine) and aminothiols (cysteamine) on E. coli cells is practically absent if the cells have genetic defects in the repair systems. This means that the explanation of radioprotection by scavenging of free radicals is invalid and that specific repair mechanisms may be involved. In order to explain the radioprotective mechanism it was suggested that the radioprotectors interact with the damaged sites in DNA so that they become partly screened from repairing endonucleases. Under these conditions the reduction of incision rate results in diminished enzymatic induction of lethal double-strand breaks in DNA, this being important only in wild type cells.To prove this hypothesis an experimental procedure was developed using bacterial cells carryng plasmids (ColE1). This procedure enabled to determine the in vivo rate of enzymatic incision of -sites. It was found that the protectors did not change the total amount of -damages in DNA but reduced the rate of enzymatic incision.     

Stability of glucose-6-phosphate dehydrogenase in human cells cultivated in vitro]

It was shown that the thermal stability of glucose-6-phosphate dehydrogenase in human diploid cells is much higher than in human heteroploid cell lines HeLa and T-9. The purified enzymes from human diploid cells and from HeLa and T-9 cells possess similar thermal stabilities. Mixing of T-9 extracts with the purified enzyme preparations revealed that the non-stability factors of the dehydrogenase are present in the T-9 extracts. An addition of NADP- and NADPH-containing buffers and crystalline NADP to the heteroploid cell extracts stabilizes the enzyme. The thermal stability of the enzyme from "in vitro" cultivated human cells depends on the concentration of the coenzyme. It was also demonstrated that glucose-6-phosphate dehydrogenase stability in HeLa and T-9 extracts is the same at low concentrations of the coenzyme and after addition of crystalline NADP. However, at NADP concentration of 10(-3) M the enzyme stability in HeLa and T-9 extracts is different. It is assumed that the destabilizing factors are the enzymes possessing the nucleotidases activity, which is different in various cell lines.