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TsOH-Catalyzed reactions of 6,7-disubstituted 2-hydroxynaphthazarins and acyclic or cyclic α,β-enones afforded the corresponding Michael-type adducts, viz., 3-substituted naphthazarins in 17–78% yields. 1H and 13C NMR and IR spectroscopy showed that the adducts with cyclohexenone and methyl vinyl ketone exist in CDCl3 solutions as mixtures of open-chain and cyclic tautomers, whereas the adducts with cyclopentenones exist only as the open-chain forms.

6,8,9-Trihydroxy-2-methyl-2H-naphtho[2,3-b]pyran-5,10-dione, a pigment of Echinothrix diadema, was synthesized. It existed as a mixture of two tautomers in CHCl3 solution according to IR spectroscopy. Its analogs 6,9-dihydroxy-2-methyl-2H-naphtho[2,3-b]pyran-5,10-dione and 6,9-dihydroxy-2-methyl-7,8-dichloro-2H-naphtho[2,3-b]pyran-5,10-dione were synthesized.

2,3-Dihydro-3-O-(1,4-naphthoquinon-2-yl)-2-oxo-1,4-naphthoquinones are the products of oxidative coupling of substituted 2-hydroxy-1,4-naphthoquinones (regardless of the presence of peri-hydroxy groups in their structures) under the action of lead dioxide.

In reaction of 5, 8-dihydroxy-2, 3-dichloro-6-ethyl-7-ethoxy-1,4-naphthoquinone and 6-tert-butyl-5,8-dihydroxy-2,3-dichloro-l,4-naphthoquinone proceeding on the surface of molecular sieves, calcium phosphate and carbonate, neutral alumina, or magnesium silicate with nucleophilic reagents (methanol, 2-methoxyethanol, monomethyl ethers of di- and triethylene glycols) activated with fluoride anion the yield of products decreased in this nucleophiles series. The most active catalysts among sorbents are alumina and magnesium silicate. All the sorbents are also efficient catalysts of alkoxy groups interchange.

It is shown that the Michael-type adducts with cyclohexanone or methyl vinyl ketone at C-3 exist as a mixture of open-chain and cyclic tautomers in CDCl3 solutions, whereas adducts with cyclopentanones at C-3 exist only in the open-chain form.

Orobol, tektorigenin, (-)-medicarpin, (±)-vestitol, and a new isoflavanone, the structure of which was established by chemical transformations and spectral data, were isolated from heartwood ofMaackia amurensis

It has been reported that rol plant oncogenes located in Ri-plasmids of Agrobacterium rhizogenes activated synthesis of secondary metabolites in the transformed plant cells. The activator mechanism is still unknown. In this work, we studied whether the NADPH oxidase-signaling pathway, which regulates the synthesis of defense metabolites in plants, is involved in the activator function of the rol genes. It was demonstrated that the transformation of Rubia cordifolia cells by the rolB and rolC genes caused an induction of biosynthesis of anthraquinone-type phytoalexins. Inhibition studies revealed a striking difference between the rolC and rolB transformed cultures in their sensitivity to Ca2+ channel blockers and calcium deficiency. The rolC culture displayed lowered resistance to the inhibitors compared to the non-transformed culture, while the rolB culture was more resistant to the treatment. The assumption was made that the oncogenic potential of rol genes is realized through the alteration of calcium balance in the plant cells. Anthraquinone production was not inhibited in the non-transformed and transformed cultures by Ca2+ channel blockers, as well as by diphenylene iodonium, an inhibitor of NADPH oxidase, and by the protein kinase inhibitor staurosporine. These results indicate that the induction of anthraquinone production in transgenic cultures does not involve the activation of Ca2+-dependent NADPH oxidase pathway.

The possibility was examined whether weak external electromagnetic radiation can affect the parameters of enzymic reactions. It was found that, in a system involving an aqueous solution of concanavalin A, glucose, and erbium salt exposed to helium-neon laser light of a wavelength of 633 nm, epimers of glucose: allosa, mannose, and galactose are formed. The effect observed was found to represent a photoinduced enzymic reaction in which the protein dissolved in water converts the glucose associated with it into epimers by the action of electromagnetic radiation. The photoepimerase activity of concanavalin A was established for the first time.

In reaction of 5, 8-dihydroxy-2, 3-dichloro-6-ethyl-7-ethoxy-1,4-naphthoquinone and 6-tert-butyl-5,8-dihydroxy-2,3-dichloro-l,4-naphthoquinone proceeding on the surface of molecular sieves, calcium phosphate and carbonate, neutral alumina, or magnesium silicate with nucleophilic reagents (methanol, 2-methoxyethanol, monomethyl ethers of di- and triethylene glycols) activated with fluoride anion the yield of products decreased in this nucleophiles series. The most active catalysts among sorbents are alumina and magnesium silicate. All the sorbents are also efficient catalysts of alkoxy groups interchange.

Interaction of the pore-forming protein (porin) from Yersinia pseudotuberculosis with S- and R-forms of the endogenous lipopolysaccharide (LPS) was studied at various ionic strengths (20-600 mM NaCl), concentrations of divalent cations (5-100 mM CaCl2, MgCl2), and pH values from 3.0 to 9.0. The interaction of the R-LPS with porin has been shown in all experimental conditions to be in consensus with the model suggesting binding at independent sites of two types. S-LPS binds to interacting sites of relatively high affinity and to independent sites of low affinity at all pH values examined and at low NaCl concentration. The cooperative interaction of the S-LPS and porin is not observed at high ionic strength and in divalent cation-free medium. The number of binding sites of porin and association constants (Ka) for both LPS forms decrease significantly on increasing the solution ionic strength. The Ka values for the R- and S-LPS change oppositely on changing the pH: the Ka value for the R-LPS is maximal (Ka = 6.7 x 10(5) M-1), but that for S-LPS is minimal (Ka = 0.4 x 10(5) M(-1) at pH 5.0-5.5. The number of high-affinity and low-affinity binding sites for both LPS forms is maximal at pH 5.0-5.5. In this case, the numbers of high- and low-affinity sites for R-LPS are 3 and 10, respectively, and those for the S-LPS are 7 and 20, respectively. These data suggest an important role of electrostatic interactions on binding of LPS to porin. The contribution of conformational changes of the ligand and protein and hydrophobic interactions are discussed.

Changes in the structure and functional activity of porin, a protein from Yersinia pseudotuberculosis, resulting from the removal of lipopolysaccharide (LPS) normally bound with the protein were studied. The treatment of LPS-containing porin with a 30% SDS solution led to an LPS-free protein that, according to the SDS-PAGE, remained to be a trimer. It was shown by CD and UV spectroscopies and intrinsic protein fluorescence that the removal of LPS caused only conformational changes in the porin secondary and tertiary structures. The LPS-free porin folded into a completely beta-structured protein aggregate. The bilayer lipid membrane technique showed that the pore-forming activity of the LPS-free porin decreased, and its concentration should be increased by two orders of magnitude to achieve the same effect. Incubation of the LPS-free porin with LPS led to a porin-LPS complex and affected the character of the protein functional activity. The treatment of the LPS-free porin by octyl glucoside, a nonionic detergent, resulted in the restoration of the protein pore-forming activity. It was suggested that the LPS and detergent provide a definite protein conformation necessary for its functioning.

The effect of solution ionic strength, calcium ion concentration, and temperature on spatial structure of cyprein was examined by CD, UV, and fluorescence spectroscopy. The secondary structure of the cyprein molecule was calculated from CD spectra, and the prevalence of the beta-structure (85%) was shown. An irreversible conformational transition in the range 55-60 degrees C was found, which reduces the binding activity of cyprein in interaction with carcinoembryonic antigen (CEA) and anti-cyprein antibodies. In the latter case, the binding activity was reversibly restored. Cyprein was shown to be a calcium-binding protein. Binding of calcium by cyprein and increasing the ionic strength of solution affect only tertiary structure of the protein. At an ionic strength of solution close to physiological conditions, calcium-bound cyprein shows maximum binding to CEA and anti-cyprein antibodies. It was shown by difference UV spectroscopy that cyprein does not interact specifically with the monosaccharides of the carbohydrate chains of CEA: fucose, mannose, galactose, and N-acetylglucosamine.

The spatial structure features of intact and deglycosylated carcino-embryonic antigen (CEA) have been studied by circular dichroism. Raman and UV-spectroscopy methods in order to elucidate a pattern and localization of CEA immunodominants. The temperature-induced changes in the spatial structure of the protein moiety were compared with data on the CEA immunochemical activity estimated by EIA procedure. A conformational transition was found within the 55-75 degrees range that produced irreversible alterations in the tertiary structure, while the secondary structure could be restored after lowering the temperature to 20 degrees C. Spectral studies of intact and deglycosylated CEA demonstrated that immunochemical activity, at least partly, was associated with the tertiary structure of the CEA protein portion.

Temperature-, ionic strength-, calcium ion- and pH-dependence of spatial structure of crustacin have been studied using CD and fluorescent spectroscopy. Secondary structure of crustacin was estimated by CD spectra. An irreversible conformational transition of crustacin's protein moiety connected with the loss of CEA-binding activity has been found at ca. 50 degrees C. Crustacin is shown to be calcium-binding protein, stability of the native crustacin conformation being markedly enhanced by calcium ions (1 mM Ca2+ shifted up the transition temperature by approximately 10 degrees C). Calcium binding and ionic strength increase led to alteration of both secondary and tertiary structures of crustacin. The highest CEA-binding activity was observed for the calcium-bond form of crustacin. A lack of specific interaction of crustacin with some saccharides was shown. Interrelation between conformation and immunochemical activity of crustacin is discussed.

Antigenic determinants of carcino-embryonic antigen (CEA) were spatially located using N-bromosuccinimide modification of tryptophan residues both in native (acetate buffer solution) and unfolded (guanidinium chloride solution) molecule of the antigen. Modification of exposed tryptophan residues failed to alter CEA antigenic activity and conformation of its protein portion as shown by CD spectroscopy. On the contrary, modification of buried tryptophan residues induced conformational changes of CEA protein portion connected with a considerable loss of its antigenic activity. It was shown that CEA antigenic activity depends on spatial structure of its protein moiety.

A glycoprotein (TSG-fs) isolated from retroplacental blood is shown to have common antigenic determinants with trophoblast-specific beta 1-glycoprotein (TSG, SP-1). Antigenic activity of TSG-fs constitute 3.4% of TSG's activity. Like TSG, glycoprotein TSG-fs contains four intramolecular disulfide bonds and belongs to proteins with predominantly beta-structure. CD-spectroscopy data give evidence of high stability of the secondary structure of TSG-fs. Immunochemical identity of TSG-fs with the reduced and carboxymethylated derivative of TSG has been demonstrated. The above data allow to suggest that TSG-fs is a fragment of the TSG molecule.

The majority of topographic antigenic determinants of the carcinoembryonic antigen (CEA) representing a glycoprotein were shown to contain sugar residues of the CEA carbohydrate chains. Periodate oxidation of CEA followed by reduction afforded a corresponding CEA derivate (CEA-POR), which retained 3% antigenic activity of CEA. In secondary and tertiary structures CEA-POR was proved to be close to CEA pre heated to 80 degrees C. Formation of borate complexes with sugar residues of CEA decreased the CEA antigenic activity to 5% while a but did not affect the spatial structure of its protein core.

Temperature- and pH-dependence of spatial structure of a native trophoblast-specific beta-glycoprotein (TSG), its desialated and deglycosylated derivatives, as well as of a fragment obtained by partial acid hydrolysis on the temperature and pH variations has been studied using CD and UV spectroscopy. Within the range 45-50 degrees C a conformational transition of the protein moiety of TSG, leading to partially reversible alterations in tertiary and secondary structures of this glycoprotein after cooling the solution to 20 degrees C has been found out. The results of spectral studies of the spatial structure of the TSG protein component have been compared with the data on antigen activity of native, temperature- and pH-denaturated, desialated, and deglycosilated TSG. It has been concluded that the protein moiety of TSG consists mainly of beta-structures; the greater part of antigen determinants of TSG is topographic and belongs to the protein component of TSG, and only 15% of antigen determinants are not connected with TSG's spatial structure.

Effects of temperature and pH on the spatial structures of trophoblast-specific beta 1-glycoprotein (TSG) and its various derivatives and fragments have been studied by circular dichroic spectroscopy. The spatial organization of the protein portion of TSG derivatives, as revealed by the spectroscopic evidence, has been discussed with respect to the antigenic activity of the species studied. We concluded that the TSG protein portion consists mainly of a beta-structural type. The antigenic determinants were shown to be topographic, and are preferentially localized in the protein portion; only about 15% of the TSG antigenic activity failed to bear a direct relation to the spatial structure. The antigenic determinants seemed to include a tryptophan residue.

Using spectroscopic methods (circular dichroism and intrinsic protein fluorescence) and immunoenzyme assay, changes in the spatial and antigenic structure of yersinin, porin from outer membrane of Yersinia pseudotuberculosis, were studied in solutions of ionic and non-ionic detergents at various temperatures and low pH values. Yersinin was shown to retain its secondary structure under various denaturation conditions, the content of regular structural patterns depending on specific action of the denaturation agent. Process of yersinin denaturation similarly to other membrane proteins appears to occur via two structural transitions: dissociation of oligomers and denaturation of monomers. At the first stage changes of quaternary structure accompanied by the loss of so called conformational determinants were observed. Temperature-dependent changes of monomers' tertiary structure affect antigenic activity of yersinin in a smaller degree.

Spatial localization of antigenic determinants of trophoblast-specific beta I-glycoprotein (TSG) has been elucidated using chemical modifications of the sugar and protein moieties of the molecule. Various deglycosylation procedures of TSG afforded fragments slightly soluble even in the presence of powerful detergents. Treatment of TSG with boric acid and its salts, accompanied with a conformational change of the sugar moiety, failed to alter conformation of the protein portion as evidenced by CD spectral data. This modification was found to increase the antigenic activity of TSG only scarcely. Modification of tryptophane or tyrosine residues of TSG changed spatial structure of the protein portion to can be a considerable loss of the TSG antigenic activity. The data obtained led to the conclusion that antigenic determinants of TSG are localized at the protein portion of the molecule and are topographic. A tryptophane residue is an indispensable constituent of the antigenic determinants.