Hydroid Obelia Longissima Research Articles

Study of the Process of Biofouling of Environmentally Safe Paint Coatings in the Natural Conditions of the White Sea

Paint coatings based on a copolymer of vinyl chloride with vinyl acetate with an additive of epoxy-diane resin of six different compositions are developed. The coatings differ in the nature of fillers (anatase, rutile, and cobalt and lanthanum oxides) and the composition of soft biocidal additives of the atrane class (protatrane of salicylic acid and hydrometallatranes based on triethanolamine complexes with zinc(II) and copper(II) salts). The process of biofouling in the natural conditions of the White sea is studied. Fouling communities formed on the studied plates during 2.5 months of exposure to the sea is analyzed, depending on the composition of the coatings. The fouling is based on the attached forms: scyphistomas (polypoid stage) of jellyfish Aurelia aurita and Cyanea sp., mussels Mytilus edulis, and hydroid Obelia longissima. The efficiency of the same paint coating appear to differ towards different species of fouling organisms.

Fluorescence properties of Ca2+-independent discharged obelin and its application prospects

Discharged obelin, a complex of coelenteramide and polypeptide, is a fluorescent protein produced from the photoprotein obelin, which is responsible for bioluminescence of the marine hydroid Obelia longissima. Discharged obelin is stable and nontoxic and its spectra are variable, and this is why it can be used as a fluorescent biomarker of variable color in vivo and in vitro. Here we examined light-induced fluorescence of Ca(2+)-independent discharged obelin (obtained without addition of Ca(2+)). Its emission and excitation spectra were analyzed under variation of the excitation wavelength (260-390 nm) and the emission wavelength (400-700 nm), as well as the 40 °C exposure time. The emission spectra obtained with excitation at 260-300 nm (tryptophan absorption region) included three peaks with maxima at 355, 498, and 660 nm, corresponding to fluorescence of tryptophan, polypeptide-bound coelenteramide, and a hypothetical indole-coelenteramide exciplex, respectively. The emission spectra obtained with excitation at 310-380 nm (coelenteramide absorption region) did not include the 660-nm maximum. The peak in the red spectral region (λ(max) = 660 nm) has not been previously reported. Exposure to 40 °C under excitation at 310-380 nm shifted the obelin fluorescence spectra to the blue, whereas excitation at 260-300 nm shifted them to the red. Hence, red emission and variation of the excitation wavelength form a basis for development of new medical techniques involving obelin as a colored biomarker. The addition of red color to the battery of known (violet to yellow) colors increases the potential of application of obelin.

Abstracts of the 17th International Symposium on Bioluminescence and Chemiluminescence ‐ (ISBC 2012)

WARNING : The light-emitting molecular structures responsible for the chemiluminescence and fluorescence phenomena are not necessarily the same!

A quantum chemical study of the formation of 2-hydroperoxy-coelenterazine in the Ca2+-regulated photoprotein obelin

The Ca2+-regulated photoprotein obelin determines the luminescence of the marine hydroid Obelia longissima. Bioluminescence is initiated by calcium and appears as a result of the oxidative decarboxylation related to the coelenterazine substrate. The luciferase of the luminescent marine coral Renilla muelleri (RM) also uses coelenterazine as a substrate. However, three proteins are involved in the in vivo bioluminescence of these animals: luciferase, green fluorescent protein, and Ca2+-regulated coelenterazine-binding protein (CBP). In fact, CBP that contains one strongly bound coelenterazine molecule is the RM luciferase substrate in the in vivo bioluminescent reaction. Coelenterazine becomes available for oxygen and the reaction with luciferase only after binding CBP with calcium ions. Unlike Ca2+-regulated photoproteins, the coelenterazine molecule is not activated by oxygen in the CBP molecule. In this work, by means of quantum chemical methods the behavior of substrates in these proteins is analyzed. It is shown that coelenterazine can form different tautomers: CLZ(2H) and CLZ(7H). The formation of 2-hydroperoxy-coelenterazine is studied. According to the obtained data, these proteins use different forms of the substrates for the reaction. In obelin, the substrate is in the CLZ(2H) form that affords hydrogen peroxide. In RM, coelenterazine is in the CLZ(7H) form, and therefore, CBP is not activated by oxygen.

Discharged photoprotein obelin: Fluorescence peculiarities

Photoprotein obelin, the enzyme–substrate complex of polypeptide with 2-hydroperoxycoelenterazine, is responsible for bioluminescence of marine hydroid Obelia longissima. Addition of Ca 2+ to the photoprotein triggers the bioluminescent reaction with light emission. The product of the bioluminescent reaction – enzyme-bound coelenteramide – is a fluorescent protein called ‘discharged’ obelin. It is stable and highly fluorescent. The paper considers dependence of its light-induced fluorescence on Ca 2+ concentration. Increase of Ca 2+ concentration enhanced the fluorescence intensity of discharged obelin; the dependence was found as linear in double logarithmic coordinates at Ca 2+ concentration range 10 −7–10 −6 М, both in excitation and emission spectra. The spectra were divided into components; contributions of the components to experimental excitation and emission spectra depended on Ca 2+ concentration. The data suggest enzymatic conformational transition in discharged obelin at ∼5 × 10 −7 M of Ca 2+ concentration. Spectra variations were attributed to acidity changes of discharged obelin chromophore (coelenteramide) in its fluorescent state S 1 ∗ .

10.1007/s11179-008-1001-8

Original and literature data supporting the evolutionary conservation of the morphofunctional organization of totipotent cells of germ and stem lineages in metazoan animals are reviewed. We studied stem cells of the colonial rhizocephalans, Peltogasterella gracilis, Polyascus polygenea and Thylacoplethus isaevae, the turbellarian Dugesia tigrina, the colonial hydroid Obelia longissima, and cultured embryonic stem cells of mouse. The typical germinal granules of germ plasm, selective expression of the activity of alkaline phosphatase and of proliferating cell nuclear antigen (PCNA), which are known as markers of stem and primary germ cells of vertebrates, and the specific expression of the protein product of the vasa gene in cells of rhizocephalans, which is a marker of cells of germ and stem lineages of various metazoans, specified the stem cells of invertebrates of such different taxa. The self-renewing pool of totipotent stem cells is the cellular basis of the reproductive strategy, including sexual and asexual reproduction; such cells share morphofunctional features of embryonic stem and germline cells of Metazoa.

Morphofunctional organization of reserve stem cells providing for asexual and sexual reproduction of invertebrates

Published and original data indicating evolutionary conservation of the morphofunctional organization of reserve stem cells providing for asexual and sexual reproduction of invertebrates are reviewed. Stem cells were studied in representatives of five animal types: archeocytes in sponge Oscarella malakhovi (Porifera), large interstitial cells in colonial hydroid Obelia longissima (Cnidaria), neoblasts in an asexual race of planarian Girardia tigrina (Platyhelmintes), stem cells in colonial rhizocephalans Peltogasterella gracilis, Polyascus polygenea, and Thylacoplethus isaevae (Arthropoda), and colonial ascidian Botryllus tuberatus (Chordata). Stem cells in animals of such diverse taxa feature the presence of germinal granules, are positive for proliferating cell nuclear antigen, demonstrate alkaline phosphatase activity (at marker of embryonic stem cells and primary germ cells in vertebrates), and rhizocephalan stem cells express the vasa-like gene (such genes are expressed in germline cells of different metazoans). The self-renewing pool of stem cells is the cellular basis of the reproductive strategy including sexual and asexual reproduction.

Spectral components of bioluminescence of aequorin and obelin

Complex bioluminescence spectra of photoproteins from marine coelenterates – jellyfish Aequorea victoria and hydroid Obelia longissima, and photoluminescence spectra of the bioluminescent reaction products (Ca 2+-discharged photoproteins) were deconvolved into components. The bioluminescence spectra of aequorin were found to include three, the bioluminescence spectra of obelin – four, and the photoluminescence spectra of the Ca 2+-discharged photoproteins – only two components. The spectral components were assigned to one unionized and three ionized forms of coelenteramide. The changes in acidity of the excited coelenteramide molecule are discussed. The differences in bioluminescence and photoluminescence spectra are considered, with protonic environment of coelenteramide taken into account.

Stem cells in reproductive strategy of asexually reproducing invertebrates

Original and literature data supporting the evolutionary conservation of the morphofunctional organization of totipotent cells of germ and stem lineages in metazoan animals are reviewed. We studied stem cells of the colonial rhizocephalans, Peltogasterella gracilis, Polyascus polygenea and Thylacoplethus isaevae, the turbellarian Dugesia tigrina, the colonial hydroid Obelia longissima, and cultured embryonic stem cells of mouse. The typical germinal granules of germ plasm, selective expression of the activity of alkaline phosphatase and of proliferating cell nuclear antigen (PCNA), which are known as markers of stem and primary germ cells of vertebrates, and the specific expression of the protein product of the vasa gene in cells of rhizocephalans, which is a marker of cells of germ and stem lineages of various metazoans, specified the stem cells of invertebrates of such different taxa. The self-renewing pool of totipotent stem cells is the cellular basis of the reproductive strategy, including sexual and asexual reproduction; such cells share morphofunctional features of embryonic stem and germline cells of Metazoa.

Bioluminescent immunoassay of thyrotropin and thyroxine using obelin as a label

Solid-phase bioluminescent immunoassay of thyroid hormones, human thyrotropin (hTSH) and two forms of thyroxine (T4), whose determinations are vitally important for diagnostics of thyroid diseases and the efficiency of treatment, is described. The recombinant obelin, a Ca 2+-regulated photoprotein originally derived from the luminous marine hydroid Obelia longissima, is employed as a bioluminescent label. To produce obelin conjugates with anti-hTSH, anti-T4 immunoglobulins (IgG), and T4, additional SH groups are introduced into the obelin molecule using Traut’s reagent (2-iminothiolane) and then obelin possessing extra SH groups is conjugated with succinimidyl 4-( N-maleimidomethyl)-cyclohexane-1-carboxylate-activated IgGs or T4. The total yield of obelin conjugates determined by luminescent activity is 60–65% after all chemical and purification procedures. The obtained conjugates are stable to lyophilization and in solution for at least 9 months at 4 °C, with loss of activity not exceeding 10%. The application of obelin conjugates for determination of the hTSH, total T4, and free T4 in standard, control, and patient sera displays high sensitivity and reproducibility of results. The results of bioluminescent immunoassays are closely comparable to those obtained by the radioimmunoassay method ( R=0.95–0.99).

Violet bioluminescence and fast kinetics from W92F obelin: structure-based proposals for the bioluminescence triggering and the identification of the emitting species.

Obelin from the hydroid Obelia longissima and aequorin are members of a subfamily of Ca(2+)-regulated photoproteins that is a part of the larger EF-hand calcium binding protein family. On the addition of Ca(2+), obelin generates a blue bioluminescence emission (lambda(max) = 485 nm) as the result of the oxidative decarboxylation of the bound substrate, coelenterazine. The W92F obelin mutant is noteworthy because of the unusually high speed with which it responds to sudden changes of [Ca(2+)] and because it emits violet light rather than blue due to a prominent band with lambda(max) = 405 nm. Increase of pH in the range from 5.5 to 8.5 and using D(2)O both diminish the contribution of the 405 nm band, indicating that excited state proton transfer is involved. Fluorescence model studies have suggested the origin of the 485 nm emission as the excited state of an anion of coelenteramide, the bioluminescence reaction product, and 405 nm from the excited neutral state. Assuming that the dimensions of the substrate binding cavity do not change during the excited state formation, a His22 residue within hydrogen bonding distance to the 6-(p-hydroxy)-phenyl group of the excited coelenteramide is a likely candidate for accepting the phenol proton to produce an ion-pair excited state, in support of recent suggestions for the bioluminescence emitting state. The proton transfer could be impeded by removal of the Trp92 H-bond, resulting in strong enhancement of a 405 nm band giving the violet color of bioluminescence. Comparative analysis of 3D structures of the wild-type (WT) and W92F obelins reveals that there are structural displacements of certain key Ca(2+)-ligating residues in the loops of the two C-terminal EF hands as well as clear differences in hydrogen bond networks in W92F. For instance, the hydrogen bond between the side-chain oxygen atom of Asp169 and the main-chain nitrogen of Arg112 binds together the incoming alpha-helix of loop III and the exiting alpha-helix of loop IV in WT, providing probably concerted changes in these EF hands on calcium binding. But this linkage is not found in W92F obelin. These differences apparently do not change the overall affinity to calcium of W92F obelin but may account for the kinetic differences between the WT and mutant obelins. From analysis of the hydrogen bond network in the coelenterazine binding cavity, it is proposed that the trigger for bioluminescence reaction in these Ca(2+)-regulated photoproteins may be a shift of the hydrogen bond donor-acceptor separations around the coelenterazine-2-hydroperoxy substrate, initiated by small spatial adjustment of the exiting alpha-helix of loop IV.

Fouling Communities of Hydrotechnical Constructions in Nakhodka Bay (Sea of Japan)

The fouling of mooring facilities in Nakhodka Bay, Sea of Japan, has been studied. The main fouling communities have been distinguished dominated by green algae Enteromorpha linza and Ulva fenestrata, brown algae Laminaria japonica and Costaria costata, a hydroid Obelia longissima, a polychaete Pseudopotamilla occelata, cirripede barnacles Balanus crenatus and Semibalanus cariosus, a bivalve mollusk Mytilus trossulus, and an ascidian Halocynthia aurantium. The naturalization of some species-invaders in the fouling of mooring facilities in Nakhodka Bay has been registered, namely hydroids Laomedea flexuosa and Clytia languida, a polychaete Pseudopotamilla occelata, and a bryozoan Bowerbankia gracilis.

Preparation and X-ray crystallographic analysis of recombinant obelin crystals diffracting to beyond 1.1 A.

Crystals of recombinant obelin, the Ca(2+)-regulated photoprotein from the marine hydroid Obelia longissima, have been grown from a solution containing PEG 8000 and potassium phosphate. Hexaminecobalt trichloride was used as an additive to increase the chance of crystallization. The crystals grow in a light yellow cubic form (0.5 x 0.5 x 0.45 mm) which diffracts to beyond 1.1 A resolution. The crystals belong to the space group C2, with unit-cell parameters a = 83.43, b = 54.92, c = 52.99 A, beta = 112.00 degrees. The asymmetric unit contains one molecule. Crystals exposed to calcium ion before and after X-ray irradiation emit light, confirming that the crystals consist of an active photoprotein.

Preparation and preliminary study of crystals of the recombinant calcium-regulated photoprotein obelin from the bioluminescent hydroid Obelia longissima.

Crystals of recombinant obelin, the Ca(2+)-regulated photoprotein from the marine hydroid Obelia longissima, have been grown from sodium citrate solutions. Crystals grow as hexagonal light-yellow rods (0.1 x 0.1 x 1.0 mm) which diffract to beyond 1.8 A with synchrotron radiation of 1.0 A wavelength. The crystals have a primitive hexagonal lattice with unit-cell parameters a = 81.55, c = 86.95 A. The asymmetric unit contains two molecules. This represents the successful preparation of single crystals of a photoprotein obelin which have promising diffraction properties.

The functional analysis of an odorant receptor using a bicistronic adenovirus vector system

Complex bioluminescence spectra of photoproteins from marine coelenterates – jellyfish Aequorea victoria and hydroid Obelia longissima, and photoluminescence spectra of the bioluminescent reaction products (Ca2+-discharged photoproteins) were deconvolved into components. The bioluminescence spectra of aequorin were found to include three, the bioluminescence spectra of obelin – four, and the photoluminescence spectra of the Ca2+-discharged photoproteins – only two components. The spectral components were assigned to one unionized and three ionized forms of coelenteramide. The changes in acidity of the excited coelenteramide molecule are discussed. The differences in bioluminescence and photoluminescence spectra are considered, with protonic environment of coelenteramide taken into account.

Positive flow dependence in the initial colonization of a fouling community: results from in situ water current manipulations

During late winter/early spring (1992–1993), a series of 12 tapered channels were placed subtidally in a Long Island, New York (U.S.A) estuary to examine total fouling community development. Rather than relying upon flow-dependent correlations among spatially segregated sites, these channels directly accelerated or decelerated the existing tidal currents ± 50% (ambient flow speeds ranged from < 2 to > 50 cm/s). In response to experimentally manipulated water flows over five months, the arborescent hydroid Obelia longissima Pallas (1766) accumulated onto Tygon tubing in direct proportion to mean free-stream flow speed, whereas barnacle ( Balanus eburneus Gould) initial settlement and cumulative recruitment along plywood ceilings increased three-fold and five-fold, respectively, with a long-term doubling of water speed. These positive flow-dependent results directly contrast those from previous experiments in the same channel apparatus which failed to document any flow enhancement on the growth of suspension-feeding bivalves.

Suspension Feeding in Oscillating Flow: The Effect of Colony Morphology and Flow Regime on Plankton Capture by the HydroidObelia longissima

The effect of flow regime on the ability of the hydroid Obelia longissima to capture plankton was examined in a laboratory flume and a wave tank. Feeding effectiveness—the proportion of the gastrozooids with tentacles successfully capturing food in a fixed amount of time—is ignificantly greater in oscillating flow than in unidirectional flow at the same average velocity and particle flux. Thus quantitative feeding studies in unidirectional flow may seriously underestimate feeding in the field if flow in the natural habitat is unsteady. Increasing colony bushiness (weight/colony length) decreases feeding effectiveness in uni-directional and low frequency oscillating flow, but not in high frequency oscillating flow. Longer colonies (0.083 m) show significantly lower feeding effectiveness than short (0.033 m) colonies. This decrease in feeding effectiveness with increasing colony length and bushiness is offset by a rapid increase in polyp number with increasing colony length.

A pelagic Isopod, Syscenus infelix, overgrown with hydroids

A specimen of the pelagic aegid isopod Syscenus infelix collected in Malangen, N. Norway, was completely overgrown with luxuriant colonies of the hydroid Obelia longissima.

The differentiation of nematocysts and associated structures in the cnidaria.

The differentiation of nematocysts and associated structures in the tentacles of the sea anemone Metridium senile fimbriatum and in the tentacles of medusae of the hydroid Obelia longissima was studied by electron microscopy. Stages in the morphogenesis of nematocysts are: 1. transformation of an interstitial cell into a cnidoblast in which the primordium of the nematocyst arises from Golgi vacuoles, 2. development of the basal part of the primordium into a capsular region that is continuous with an external tube with which microtubules, Golgi apparatus, and centrioles are associated distally, 3. growth of the capsular region to include most of the external tube, and 4. differentiation of an internal tube with spines from the matrix of the capsule and of the remaining external tube. The external tube is not invaginated to become the internal tube, but is incorporated into the capsular region by distal growth of the capsular wall. At the same time the matrix, both before and during differentiation into a thread, moves into the capsule. The development of nematocysts in Obelia differs from that in Metridium in several respects. In the former the basitrich has an operculum plus associated cnidocil and dense rods whereas in the latter the nematocysts (amastigophore, b-mastigophore and basitrich) have three apical flaps and an associated flagellum, with the exception of the spirocyst.