[The mechanism of formation of emitter by fluorescence of calcium-discharged obelin].

It is known that bioluminescence of obelin is triggered by Ca2+ the binding of which to the protein induces the decarboxylation of 2-hydroperoxycoelenterazine. The molecular mechanism of fluorescence of obelin, which determines the fluorescence of see hydroid Obelia Longissima, has been investigated with the use of quantum chemical calculations. According to quantum chemical calculations, the emitter of the reaction is the ion-pair state of phenolate-anion of coelenteramide. It has been shown that the fluorescence spectrum of this state depends on the position of the proton between the oxygen atoms of the phenol group of coelenteramide and the nitrogen atom of His22. The agreement of the calculated absorption and fluorescence spectra with the experimental spectrum shows the accuracy of the quantum chemical calculations and conclusions.

[Calcium-regulated photoproteins of marine coelenterates].

Ca(2+)-regulated photoproteins are bioluminescent proteins responsible for bioluminescence of marine coelenterates. The photoprotein molecule is a stable enzyme-substrate complex consisting of a single polypeptide chain and an oxygen "pre-activated" substrate, 2-hydroperoxycoelenterazine, which is tightly but non-covalently bound with a protein. The bioluminescence is triggered by calcium ions and originates from an oxidative decarboxylation of a protein bound substrate. The review provides current data on the photoproteins structure, the mechanism of bioluminescent reaction, the function of some amino acid residues of an active site in the catalysis and the formation of the emitter, as well as on applications of these proteins in a bioluminescent analysis.

[Conjugates of the Ca2+ -regulated photoprotein obelin with immunoglobulins: synthesis and use as labels in bioluminescent immunoassay]. / Sintez kon''iugatov Ca-reguliruemogo fotoproteina obelina s immunoglobulinami i ikh ispol'zovanie v kachestve metok v immunoanalize.

An efficient procedure for obelin conjugation with immunoglobulins was developed. The possibility was shown of using the resulting conjugates instead of a radioisotope label for the immunoassay of thyroid stimulating hormone in sera; the conjugates provide a sensitivity of 0.01 microIU/ml. The results of bioluminescent immunoassay (sera of 34 patients) satisfactorily correlate with the results of radioisotope assay (R 0.99).

[The Ca2+-activated photoprotein obelin as a calcium transport inducer in proteoliposomes in the membrane of the T-system of skeletal muscles]. / Ca2+-aktiviruemyi fotoprotein obelin kak indikator kal'tsievogo transporta v proteoliposomakh, vkliuchaiushchikh membrany T-sistemy skeletnykh myshts.

The calcium-binding photoprotein obelin extracted and purified from the luminescent hydroid Obelia longissima was used to record the processes of Ca2+ release from proteoliposomes. It has been shown that lecithin proteoliposomes with incorporated rabbit skeletal muscle T-system membranes possess a BAY K-8644-activated permeability which is inhibited by nitrendipine. The Ca(2+)-activated photoprotein obelin is a convenient and perspective tool in studies of fast calcium fluxes.

[NADPH- and ATP-dependent luminescence of extracts from luminous bacteria]. / Zavisimaia ot NADPH i ATP liuminestsentsiia ékstraktov svetiashchikhsia bakterii.

It was shown that the luminescence of extracts prepared from luminous bacteria is stimulated by NADPH and ATP without FMN or long-chain aliphatic aldehydes, which are routinely used for producing luminescence of extracts from luminous bacteria in vitro. In these extracts an aldehyde factor, a natural analog of aliphatic aldehydes, is synthesized. The enzymatic system involved in maintaining the luminescence of NADPH and ATP is probably not coupled with the functioning of NAD(P)H: FMN oxidoreductase which has been supposed to participate in luminescence processes in vivo. It is assumed that both aldehyde factor synthesis and reduction of endogenous analog of FMN, natural substrates of bacterial luciferase, are due to the functioning of the same metabolic pathway.

[Effect of phenobarbital on the luminescence system of luminous bacteria]. / Vliianie fenobarbitala na liuminestsentnuiu sistemu svetiashchikhsia bakterii.

The effect of phenobarbital on the luminescent system of Beneckea harveyi was studied. The inhibition of luminescence with phenobarbital was shown to be due to a disorder in the synthesis of an aldehyde factor, the endogenous substrate of bacterial luciferase. Upon the action of phenobarbital, the bacterium acquires the properties of "aldehyde" mutants, i. e. their luminescence is stimulated with exogenous decyl aldehyde. The luminescence of the cells was also stimulated with long-chain aldehydes, fatty acids and their analogues: apparently, the aldehyde factor is formed via incorporation of an oxygen atom into the terminal methyl of a saturated fatty acid or its analogue. Phenobarbital has no effect on the bacterial growth; however, it increases the content of luciferase in the culture. The results suggest that phenobarbital is not a direct inductor of luciferase synthesis. Possibly, the stimulating action of phenobarbital involves the inhibition of synthesis of the aldehyde factor and, consequently, an increase in the concentration of intermediate products of its synthesis.

[Lipids of the luminescent bacteria Photobacterium mandapamensis]. / Lipidy svetiashchikhsia bakterii Photobacterium mandapamensis.

The composition of lipids was studied in the luminescent bacterium Photobacterium mandapamensis under the conditions of maximal luminescence. The synthesis of total lipids and poly-beta-hydroxybutyric acid (PHBA) was investigated in dynamics under the conditions of batch cultivation. The major class of lipids was polar lipids (84.3%) represented by phospholipids (phosphatidyl ethanolamine, phosphatidyl glycerol, cardiolipin, lysocardiolipin, lysophosphatidyl ethanolamine) and a minor nonidentified phospholipid. The fraction of neutral lipids was represented mainly by free fatty acids (about 5%), tri- and diglycerides (in trace amounts), and two nonidentified classes, apparently, hydrocarbons and waxes. A correlation was established between the luminescence of the bacterium and the content of PHBA.

[Fatty acid makeup changes in the luminescent bacteria, Photobacterium mandapamensis, in periodic cultivation]. / Izmenenie zhirnokislotnogo sostava svetiashchikhsia bakterii Photobacterium mandapamensis pri periodicheskom kul'tivirovanii.

The fatty acid composition of the luminescent bacterium Photobacterium mandapamensis and its spontaneous dark mutant was studied in dynamics. Lipids of the both strains extracted with a methanol -- chloroform mixture contained the following fatty acids: lauric, tridecanoic, myristic, tetradecenic, pentadecanoic, pentadecenic, palmitic, palmitoleic, heptadecanoic, C17-cyclopropanoic, stearic, octadecenic, and nonadecanoic. The content of palmitoleic acid was the highest (57% of the total). Not all of the acids changed their content in the same direction during batch cultivation of the luminescent and dark strains. The content of palmitoleic acid fell to 49.2% of the total in the luminescent culture at the point of its maximal luminescence, but it increased to 63.8% in the dark strain at the corresponding growth phase.

[Luciferase synthesis regulation in Photobacterium mandapamensis]. / Reguliatsiia sinteza liutsiferazy u Photobacterium mandapamensis.

The synthesis of luciferase and the dynamics of luminescence were studied in the course of batch cultivation of the strain 54-K obtained from the wild strain of Photobacterium mandapamensis after numerous passages. Luciferase synthesis bvy the strain was not sensitive to the inhibitor contained in the growth medium and did not require the accumulation of an "audoinductor". Since the intensity of luminescence and the content of luciferase per cell did change, the bacterium seemed to possess an additional system for regulating the synthesis of luciferase which was independent of the "autoinductor" and the inhibitor.

[Luminescence and growth of Photobacterium mandapamensis in periodic culture]. / Liuminestsentsiia i rost Photobacterium mandapamensis v periodicheskoi kul'ture.

The luminescent Photobacterium mandapamensis, strain 54 (the collection of the Institute of Physics, Siberian Branch of the USSR Academy of Sciences), was isolated from the water of the Pacific Ocean in the equatorial zone and studied in the course of periodic cultivation. The dynamics of changes in the main parameters of the culture, such as luminescence, growth, respiration, heat emission etc., was investigated. The data obtained were used to establish a correlation between changes in these parameters and in the cell metabolism when one energy substrate was substituted by another.