Characterization of two glutamate dehydrogenases from the symbiotic microalga Symbiodinium microadriaticum isolated from the coral Acropora formosa

In view of their possible involvement in ammonium assimilation in the coral/algal symbiosis, we have purified two distinct glutamate dehydrogenase isoenzymes from the symbiotic dinoflagellate Symbiodinium microadriaticum (Freudenthal) extracted from the staghorn coral Acropora formosa collected from Magnetic Island, North Queensland, Australia, in 1986–1987. An NADPH-specific glutamate dehydrogenase (GDH) displayed biphasic kinetics with respect to ammonium as the variable substrate; at low substrate concentrations the apparent K _m was below 1 mM, whereas at high substrate concentrations the corresponding value was approximately 200 mM. The NADPH-GDH displayed extremely low activity in the direction of glutamate oxidation; together with the kinetic data this suggests a probable role in ammonium assimilation. A second (NADH-specific) GDH was found to have both amination and deamination activities, and presumably functions in vivo in glutamate oxidation. Kinetic constants are reported for both GDH isoenzymes.     

NADP+-isocitrat-dehydrogenase aus Idus idus (Pisces: Cyprinidae). II. Einfluß der temperatur auf substrat- und cosubstrataffinität

Maximum substrate and cosubstrate affinity, as judged by the Michaelis constant (K _M ), of NADP⁺-dependent isocitrate dehydrogenase of pig heart (purchased from Boehringer, Mannheim, FRG) is attained at 37°C. If K _M -values of substrate (Isocitrate, IC) and cosubstrate (NADP⁺) of NADP⁺-dependent isocitrate dehydrogenase (ICDH) of the white dorsal muscle of Idus idus L. is plotted against the experimental temperature (VT), W-shaped curves result. With increasing adaptation temperature (AT), there is a shift to increasing VT. It is suggested that the W-shaped curves are due either to the simultaneous presence of two multiple forms of the enzyme, or to the reversible temperature-dependent interconversion of one protein species.     

Role of glutamine synthetase in ammonia assimilation by symbiotic marine dinoflagellates (zooxanthellae)

The dinoflagellate symbionts (zooxanthellae) present in many reef corals aid in the survival of the symbiotic unit in nitrogen deficient tropical waters by providing additional routes of nitrogen uptake and metabolism. The enzymatic pathway of ammonia assimilation from seawater and the re-assimilation of coral ammonium waste by zooxanthellae was studied by examining the affinity of glutamine synthetase for one of its substrates, ammonia. Glutamine synthetase activity was measured in dinoflagellates of the species Symbiodinium microadriaticum found in symbiotic association with various marine coelenterates. Michaelis-Menten kinetics for the substrate ammonia were determined for freshly isolated dinoflagellates from Condylactis gigantea (apparent NH₃ K_m=33 M) and for cultured dinoflagellates from Zoanthus sociatus (apparent NH₃ K_m=60 M). On the basis of the low apparent K_ms for NH₃, it appears that ammonia assimilation by these symbiotic dinoflagellates occurs via the glutamine synthetase/glutamate synthase pathway. Additionally, the uptake of exogenous ammonium by an intact coelenterate-dinoflagellate symbiosis was strongly inhibited by 0.5 mM methionine sulfoximine, and inhibitor of glutamine synthetase.     

The ammonium/methylammonium uptake system of Symbiodinium microadriaticum

The substrate analogue [¹⁴C]-methylammonium was used to study ammonium/methylammonium uptake by Symbiodinium microadriaticum (zooxanthellae). The value of the Michaelis constant (K _m) for the uptake system was approximately 35 M with methylammonium as substrate; ammonium was a competitive inhibitor of methylammonium uptake, and the K _m for ammonium uptake (determined as the inhibition constant, K _i, for methylammonium) was 6.6 M. Methylammonium uptake by zooxanthellae was light-dependent. Methylammonium uptake rates of zooxanthellae which had been freshly isolated from the hermatypic coral Acropora formosa (0.85±0.05x10^-10 mol min^-1 cell^-1) were lower than those of axenic cultures of the zooxanthellae from Montipora verrucosa (Acroporidae) grown under various nitrogen regimes (1.6 to 12x10^-10 mol min^-1 cell^-1). Maximum uptake rates were found for ammonium-starved cultured M. verrucosa zooxanthellae (10.2 to 12x10^-10 mol min^-1 cell^-1); M. verrucosa zooxanthellae growing with ammonium as nitrogen source and zooxanthellae which had been freshly isolated from A. formosa gave similar and considerably lower uptake rates (0.85 to 1.6x10^-1 mol min^-1 cell^-1). These results suggest that either coral tissue contains sufficient ammonium to repress synthesis of the uptake system of the algal symbionts or, alternatively, there are additional barriers to ammonium transport for zooxanthellae in vivo.     

Metabolic enzymes in coelomic cells (eleocytes) of the polychaete Nereis virens: sex specific changes during sexual maturation

The activities of some enzymes of the intermediary metabolism and the content of soluble protein and carbohydrate (glycogen plus free glucose) were measured in one type of coelomic cells (eleocytes) of the polychaete Nereis virens. Specimens used in this study were collected between 1989 and 1991 in Oosterscheldt Bay, The Netherlands, and divided into six different stages of sexual maturation as determined by the mean oocyte volume. In both sexes, the soluble protein content in eleocytes of immature individuals (11 mg ml^–1 cell vol) increased three-fold. In prespawning N. virens the soluble protein content decreased to less than 2 mg protein ml^–1 cell vol in females but not in males. In both sexes, the carbohydrate content decreased continuously from immature [300 mol glucose equivalent (equiv) ml^–1 cell vol] to prespawning individuals (< 40 mol glucose equiv ml^–1 cell vol). During the time course of maturation, the specific activities (expressed as units mg^–1 protein) of pyruvate kinase, phosphoenolpyruvate carboxykinase, malate dehydrogenase, alanine aminotransferase and glutamate dehydrogenase decreased in both sexes. A transient increase in the specific activities was found for glycogen phosphorylase and aspartate aminotransferase. No major changes were found for hexokinase, lactase dehydrogenase, glucose-6-phosphate dehydrogenase and malic enzyme. Sex specific differences were found for the activities of citrate synthase and isocitrate dehydrogenase, which were higher in males. the specific activities of the latter enzyme increased more than ten-fold in males, but only four-fold in female eleocytes during maturation. In eleocytes of prespawning females, the activities of most enzymes showed extremely high variations not found in prespawning males. For two enzymes of fatty acid catabolism, -hydroxyacyl-CoA dehydrogenase and -hydroxybutyrate dehydrogenase, only traces of activities were detected, suggesting the absence of significant fatty acid catabolism in the eleocytes. Compared to the eleocytes, the body wall tissue showed ten-fold higher activities of phosphofructokinase, whereas the eleocytes displayed higher activities of the amino acid interconverting enzymes glutamate dehydrogenase and alanine aminotransferase and the glyconeogenic enzyme phosphoenolpyruvate carboxykinase. Citrate synthase activities were similar for both tissues. In the coelomic fluid of N. virens, glucose (< 0.1 to 3.5 mM) and d-lactate (0.1 to 4 mM) were present and represent exogenous substrates for the eleocyte metabolism.     

Increased mortality and photoinhibition in the symbiotic dinoflagellates of the Indo–Pacific coral Stylophora pistillata (Esper) after summer bleaching

Coral bleaching (the loss of symbiotic dinoflagellates from reef-building corals) is most frequently caused by high-light and temperature conditions. We exposed the explants of the hermatypic coral Stylophora pistillata to four combinations of light and temperature in late spring and also in late summer. During mid-summer, two NOAA bleaching warnings were issued for Heron Island reef (Southern Great Barrier Reef, Australia) when sea temperature exceeded the NOAA bleaching threshold, and a ‘mild’ (in terms of the whole coral community) bleaching event occurred, resulting in widespread S. pistillata bleaching and mortality. Symbiotic dinoflagellate biomass decreased by more than half from late spring to late summer (from 2.5×10⁶ to 0.8×10⁶ dinoflagellates cm² coral tissue), and those dinoflagellates that remained after summer became photoinhibited more readily (dark-adapted F _V : F _M decreased to (0.3 compared with 0.4 in spring), and died in greater numbers (up to 17% dinoflagellate mortality compared with 5% in the spring) when exposed to artificially elevated light and temperature. Adding exogenous antioxidants (d-mannitol and l-ascorbic acid) to the water surrounding the coral had no clear effect on either photoinhibition or symbiont mortality. These data show that light and temperature stress cause mortality of the dinoflagellate symbionts within the coral, and that susceptibility to light and temperature stress is strongly related to coral condition. Photoinhibitory mechanisms are clearly involved, and will increase through a positive feedback mechanism: symbiont loss promotes further symbiont loss as the light microenvironment becomes progressively harsher.     

Purification and partial characterisation of phenoloxidase from the colonial ascidian Botryllus schlosseri

Phenoloxidase (PO) from the colonial ascidian Botryllus schlosseri was purified using two different chromatographic strategies. A three-step purification was developed in order to maintain enzyme activity, whereas an easier purification procedure was adopted to obtain enough PO for the production of specific polyclonal antibodies. The enzyme showed optimal pH and temperature values of 7.0 to 7.5 and 35 °C, respectively, and a K _m value of 4.62 ± 0.76 mM was estimated using l-DOPA as substrate. A molecular weight of 160 kDa was determined after SDS-PAGE under non-reducing conditions. The addition of the reducing agent β-mercaptoethanol caused the disappearance of the 160 kDa band and the appearance of a new band at 80 kDa, suggesting that active PO is a dimer and the two subunits are linked by disulphide bridges. Received: 14 December 1998 / Accepted: 24 August 1999     

Nitrogen excretion by some demersal macrozooplankton in Heron and One Tree Reefs,Great Barrier Reef,Australia

Nitrogen excretion rates of demersal macrozooplankton were measured together with nitrogen concentrations in the water column and sediments in lagoons of Heron Reef and One Tree Reef, Great Barrier Reef, Australia, during August and November 1991. Excretion rates increased with body weight, and weight-specific excretion rates of the demersal macrozooplankton were comparable to those of pelagic zooplankton and meiofauna in the Great Barrier Reef. Values of demersal macrozooplankton abundance from previous studies and excretion rates from this study were combined to estimate fluxes of ammonium from demersal macrozooplankton in coral reef lagoons. The estimated fluxes in the water column and sediments were 12 M NH₄ m^-2 d^-1 and 34 M NH₄ m^-2d^-1, respectively. These fluxes were compared with reported fluxes of ammonium in coral reef lagoons in the Great Barrier Reef, Australia. The estimated flux from the demersal macrozooplankton in the water column was 29 and 9% of those reported for microheterotroph regeneration and phytoplankton utilization, respectively. It was 10% of the reported advective flux during periods of low advection and 13% of the maximum efflux from sediments computed from diffusion models. The estimated flux from the demersal macrozooplankton in the sediments exceeded those reported for meiofauna, and was 5 to 32% and 2 to 13% of those reported for ammonification and utilization in sediments, respectively. The potential importance of demersal macrozooplankton in mediating sediment-water column exchanges in the absence of diffusive effluxes and when they swarm is discussed.     

A comparative study of the glutamate dehydrogenase activity in several species of marine phytoplankton

Kinetic studies on the NADP-linked glutamate dehydrogenase (GDH) activity in 8 species of marine phytoplankton have been carried out. The pH optima, linearity of reactions, apparent energies of activation, and apparent K _m values for NADPH, NH ₄ ^>+ and -ketoglutarate were determined. Based on these data, an accurate, sensitive enzyme assay procedure is presented which was successfully tested on shipboard during a recent cruise in an upwelling region. Preliminary studies on inhibition and activation of the NADP-linked GDH activity have also been carried out. In addition to NADP-linked GDH activity, NAD-linked GDH activity was discovered in certain species of phytoplankton. The possible physiological roles of NAD-and NADP-linked GDH enzymes are discussed.Contribution No. 911 from the Department of Oceanography, University of Washington. This research was supported by the National Science Foundation Grants GA-34165 and GX-33502.     

Metabolic specialization of mitochondria from scallop phasic muscles

In fast, glycolytic muscles, oxidative phosphorylation presumably facilitates recuperation from exhaustive exercise and supports growth and maintenance metabolism. Given the shifts in pH with extensive glycolytic activity, the pH optima of mitochondrial processes should indicate whether mitochondria are adapted for recuperation from exercise or for growth and maintenance. We examined this question using mitochondria from the phasic adductor muscle of the scallop, Euvola (Pecten) ziczac, collected from the Golfo de Cariaco, Venezuela in 1992 and 1993. Scallop muscle mitochondria showed well coupled oxidation of glutamate and pyruvate at pH 7.0 and 6.4. The preferred substrates (glutamate, pyruvate and succinate) were oxidized at approximately 40 nmol O₂ min^-1 mg^-1 mitochondrial protein at 25°C, while malate and glutamine were oxidized at 75% and proline at 30% of these rates. Neither palmitoyl carnitine nor aspartate were oxidized. Succinate oxidation was not coupled to ADP utilization at pH 7.0 but was somewhat coupled at pH 6.4. Generally, State 3 rates of oxygen uptake were similar at pH 7.0 and 6.4. Maximal rates of oxidation of glutamate and pyruvate showed broad pH optima. For both glutamate and pyruvate, the highest respiratory control ratio (RCR) values were found at pH 6.5. The saturation curves of scallop muscle mitochondria for pyruvate, glutamate and ADP were well described by the Michaelis-Menten equation. The affinity for pyruvate was greater at pH 6.4 (apparent K _{m, app}=0.013 mM) than at pH 7.0 (K _{m, app}=0.026 mM) while the affinity for ADP (K _{m, app}=0.015 mM) and that for glutamate (K _{m, app}=0.55 mM) changed little with pH. The ADP affinity was the same whether pyruvate or glutamate was the carbon substrate. The combination of maintenance of sensitivity to ADP with an enhanced affinity for pyruvate at acidic pH values should facilitate recuperation from bouts of glycolytic activity. Scallops harvested in September and those harvested in January differed in the maximal rates of glutamate and pyruvate oxidation.     

Effects of methionine sulfoximine on growth and nitrogen assimilation of the marine microalga Chlorella autotrophica

Chlorella autotrophica Shihira and Krauss (clone 580), a euryhaline microalga from the marine coastal environment is subject to large fluctuations in external salinity and nitrogen supply. The alga exhibits maximum growth at salinities lower than 100% ASW (artificial seawater). Cells divide faster and show higher cell yields when the supply of either NH ₄ ⁺ or NO ₃ ^- is increased above 0.2 mM. Cells growing on NH ₄ ⁺ show high levels of NADPH-glutamate dehydrogenase (GDH) activity, and the levels of glutamine synthetase (GS) are decreased to very low levels under these conditions. Methionine sulfoximine (MSX), an inhibitor of GS, has little effect on cell division and nitrogen assimilation of cells growing on NH ₄ ⁺ . Cells growing on NO ₃ ^- , however, show marked inhibition (65%) in nitrogen assimilation in the presence of 5 mM MSX. This MSX concentration also causes growth retardation and a progressive decrease in cell protein and nitrogen content. GS is almost completely inhibited by 5 mM MSX in both NH ₄ ⁺ and NO ₃ ^- -grown cells. Cells growing on NH ₄ ⁺ maintain high levels of NADPH-GDH activity in the presence of MSX. NADPH-GDH activity in MSX-treated NO ₃ ^- -grown cells increases, and, in the presence of 5 mM MSX, reaches 40% of the level found in NH ₄ ⁺ -grown cells. These results are consistent with NADPH-GDH providing an alternate pathway for NH ₄ ⁺ assimilation by this marine Chlorella species.     

Purification and characterisation of octopine dehydrogenase from the marine nemertean Cerebratulus lacteus (Anopla: Heteronemerta): comparison with scallop octopine dehydrogenase

Octopine dehydrogenase from the nemertean Cerebratulus lacteus was purified over 1000-fold to almost homogeneity. The enzyme does not bind to arginine Sepharose 4B. It has a monomeric structure with a relative molecular mass of 40000. Two isoenzymes were identified with isoelectric points of 5.6 and 5.4, whereas the purified isoenzymes of Pecten jacobaeus adductor mucles (which bind to arginine Sepharose 4B) had lower IEP's of 4.9 and 4.7. Apparent K_m's of the nemertean ODH for arginine and pyruvate are dependent on the respective co-substrate concentration. This phenomenon may result in activation of ODH and, thus, production of octopine in locomotory highly active individuals while attacking food, especially when this takes place in a hypoxic habitat, such as decaying mud near the high-water mark. The apparent K_m's for octopine (0.22 mM) and NAD⁺ (14 M) are low. Octopine is a substrate inhibitor for the reverse reaction above 2 mM, and a product inhibitor of the forward reaction by 50% at 1.2 mM. Therefore, only small amounts of octopine are likely to accumulate in vivo. Amino acid substrate specificity is limited to guanidino amino acids. We believe that the amino acid substrate specificity is not an evolutionary modification, but rather that it is narrowed to guanidino amino acids (or even specificity to arginine) in those species where ODH has a physiological function in maintaining redox balance during exercise. The specificity for keto acids is dependent on chain length, (-ketobutyrate>-ketocapronate); a second carboxyl group inactivates the enzyme.     

Glycerol translocation in Condylactis gigantea

Sodium cyanide (NaCN) was used to partially uncouple respiration and photosynthesis in the symbiotic sea anemone Condylactis gigantea. NaCN significantly increased the ratio of gross photosynthesis to respiration in both intact tentacles and isolated zooxanthellae (Symbiodinium microadriaticum), increased carbon translocation from 17.7±3.5% of total fixed in controls to 43.5±5.8%, and doubled the amount of photosynthetically fixed carbon accumulating in the coelenterate host over that in controls. Only 2% of the non-particulate radioactivity recovered in the host tissue was ¹⁴C-glycerol when uninhibited symbiotic tentacles were incubated in ¹⁴C-bicarbonate for 1 h. At 10^-5 M NaCN, approximately 25% of the host nonparticulate radioactivity was recovered as ¹⁴C-glycerol, the absolute concentration of glycerol in the host tissue was three times higher than in controls, and ¹⁴C-glycerol was found in the medium. While glycerol has been proposed to play a major role in the translocation of photosynthetically fixed carbon from zooxanthellae to their coelenterate hosts, its concentration has never been measured in the animal and algal components of the symbiosis. The isolated zooxanthellae contained 3.62±0.33 mM glycerol, 26x the 0.141±0.02 mM found in the anemone. Aposymbiotic anemone tissue contained 0.169±0.06 mM glycerol. The rate of glycerol mineralization was not saturated even when exogenous glycerol levels were 70x internal concentrations. These data show that respiration and photosynthesis in symbiotic associations may be partially uncoupled by NaCN, and that this uncoupling allows the verification of the translocation and rapid catabolism of glycerol within the host.     

Adenylate cyclase of Mytilus galloprovincialis posterior adductor muscle: basal properties and sensitivity to serotonin

The presence and characteristics of a membrane-bound adenylate cyclase from Mytilus galloprovincialis Lmk posterior adductor muscle have been investigated. The enzyme has a Michaelis constant (K _m) of 0.38 mM at 20°C and requires divalent cations (Mg²⁺/Mn²⁺) for its activity. Optimal GTP concentrations are between 10^-5 and 10^-4 M. The non-hydrolizable GTP-analogues GMPpNHp and GTPS increase the activity of the enzyme four- to ten-fold. Sodium fluoride stimulates the enzymatic activity seven- to eight-fold. Forskolin increases the enzymatic activity two- to three-fold. Serotonin stimulates the adenylate cyclase activity in a dose-dependent manner. These experiments were performed with mussels collected from the estuary of Muros, Spain, in 1990.     

A new growth model for fishes and the estimation of optimum age of fish populations

Evidence is presented that a simple power equation of the formX _t –X _m =±b(|tࢤt _m|)^B can describe growth in length and weight of fishes, whereX _t denotes fish length or weight at aget, X _m is length or weight (L _m orW _m ) at a reference aget _m , andb andB are parameters to be estimated by the least squares. The optimum age of fish populations (t _y ) may be estimated by the equationMW _m /b=±y ^B (B/y-M) whereM denotes natural mortality and wherey=t _y -t _m .     

Molecular mechanisms of adaptation to low temperature in marine poikilotherms. Some regulatory properties of dehydrogenases from two arctic species

The properties of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase from gill tissue of the tanner crab Chionocetes bairdi, and lactate dehydrogenase (LDH) and glyceraldehyde dehydrogenase from skeletal muscle of C. bairdi and the yellowfin sole Limanda aspera were examined over the physiological temperature range of the animals. Both animals were obtained in the Bering Sea in winter, and their enzymes appear to be remarkably cold-adapted. Affinity of sole LDH for substrate appears to increase with decreasing temperature, thus keeping reaction rate essentially independent of temperature at physiological concentrations of the substrate. Calculated values of activation energy are low, in keeping with the argument that organisms from cold environments have enzymes with a reduced energy of activation. In addition, Hill plots of the substrate saturation curves for lactate dehydrogenase from muscle of sole indicate that there is a facilitation of allosteric behaviour at low temperatures. Maximum affinity of sole LDH for substrate in the absence of univalent cations occurs at 3°C, while in the presence of 150 mN K⁺, it occurs between 0° to-2°C. The effects of Mg²⁺ on enzyme activity appear to be determined by concentration of substrate and temperature. Thus, glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase are stimulated more effectively by Mg²⁺ at low temperature and at low substrate levels whereas, at high concentrations of substrate, they are relatively independent of the bivalent cation. All four dehydrogenases are affected by the univalent cations Na⁺, K⁺ and NH₄ ⁺ in a manner which appears to be determined, in part at least, by concentration of substrate and by temperature. These findings suggest mechanisms for the maintenance and regulation of enzyme activity in poikilothermic tissues at low and changing temperatures.     

Sediment ammonium availability and eelgrass (Zostera marina) growth

The interaction of sediment ammonium (NH ₄ ⁺ ) availability and eelgrass (Zostera marina L.) growth, biomass and photosynthesis was investigated using controlled environment and in-situ manipulations of pore water ammonium concentrations. Sediment diffusers were used to create pore water diffusion gradients to fertilize and deplete ammonium levels in sediments with intact eelgrass rhizospheres. Between October, 1982 and September, 1983 controlled environment experiments using plants from shallow (1.3 m) and deep (5.5 m) stations in a Great Harbor, Woods Hole, Massachusetts, USA eelgrass meadow along with in-situ experiments at these stations provided a range of sediment ammonium concentrations between 0.1 and 10 mM (adsorbed+interstitial NH ₄ ⁺ ). The results of the in-situ experiments indicate that nitrogen limitation of eelgrass growth does not occur in the Great Harbor eelgrass meadow. A comparison of NH ₄ ⁺ regeneration rates and eelgrass nitrogen requirements indicates an excess of nitrogen supply over demand and provides an explanation for the lack of response to the manipulations. Results of controlled environment experiments combined with in-situ results suggest that sediment ammonium pool concentrations above approximately 100 mol NH ₄ ⁺ per liter of sediment (interstitial only) saturate the growth response of Zostera marina.     

Effects of nitrogen and phosphorus enrichement on in vivo symbiotic zooxanthellae of Pocillopora damicornis

The reef coral Pocillopora damicornis (Linnaeus) was grown for 8 wk in four nutrient treatments: control, consisting of ambient, unfiltered Kaneohe Bay seawater [dissolved inorganic nitrogen (DIN, 1.0 M) and dissolved inorganic phosphate (DIP, 0.3 M)]; nitrogen enrichment (15 M DIN as ammonium); phosphorus enrichment (1.2 M DIP as inorganic phosphate); and 15 M DIN+1.2 M DIP. Analyses of zooxanthellae for C, N, P and chlorophyll a after the 8 wk experiment indicated that DIN enrichment increased the cellular chlorophyll a and excess nitrogen fraction of the algae, but did not affect C cell^-1. DIP enrichment decreased both C and P cell^-1, but the decrease was proportionally less for C cell^-1. the response of cellular P to both DIN and DIP enrichment appeared to be in the same direction and could not be explained as a primary effect of external nutrient enrichment. The observed response of cellular P might be a consequence of in situ CO₂ limitation. DIN enrichment could increase the CO₂ (aq) demand by increasing the net production per unit area. DIP enrichment could slow down calcification, thus decreasing the availability of CO₂ (aq) in the coral tissue.Hawaii Institute of Marine Biology Contribution No. 920     

Photosynthetic carbon acquisition in the red algaGracilaria conferta

Photosynthetic properties of the common red algaGracilaria conferta, collected from the eastern Mediterranean Sea were investigated in 1989, in order to begin evaluating its adaptative strategies with regard to the inorganic carbon composition of seawater, and to test whether the alleged C₄ photosynthesis of anotherGracilaria species is common within the genus. Net photosynthetic rates ofG. conferta were, under ambient conditions of inorganic carbon (ca. 10µM, CO₂ and 2.2 mM HCO ₃ ^- ), not sensitive to O₂ over the range 10 to 300µM, and the CO₂ compensation point was low (ca. 0.005µM). Ribulose-1,5-bisphosphate carboxylase/oxygenase was the major carboxylating enzyme, with a crude extract activity of 175µmol CO₂ g^–1 fresh wt h^–1 while phosphoenolpyruvate carboxylase and phosphoenolpyruvate carboxykinase were present at 70 and 20%, respectively, of that activity. No activities of the decarboxylases NAD-and NADP-malic enzyme could be detected. The¹⁴C pulse-chase incorporation pattern showed thatG. conferta fixes inorganic carbon via the photosynthetic carbon reduction cycle only, with no evidence for photosynthetic C₄ acid metabolism. Photosynthesis at the natural seawater pH of 8.2 was, at 25°C and saturating light, saturated at the ambient inorganic carbon concentration of 2.5 mM. It is proposed that, under ambient inorganic carbon conditions, a CO₂ concentrating system other than C₄ metabolism provides an internal CO₂ concentration sufficient to suppress the O₂ effect on ribulose-1,5-bisphosphate carboxylase/oxygenase and, thus, on photorespiration, in a medium where the external free CO₂ concentration is lower than theK _m(CO₂) of the carboxylating enzyme. Since inorganic carbon, under natural saturating light conditions, seems not to be a limiting factor for photosynthesis ofG. conferta, it likely follows that other nutrients limit the growth of this alga in nature.     

NADP+-isocitrat-dehydrogenase aus Idus idus (Pisces: Cyprinidae). I. Aktivität als funktion der adaptationstemperatur

Activity of NADP⁺-dependent isocitrate dehydrogenase (ICDH) of the white dorsal muscle of Idus idus L. was determined by special analysis, under optimum homogenisation, centrifugation and cuvette conditions. The experimental conditions differed considerably from those used for the investigation of NADP⁺-ICDH activity in mammals by Bücher et al. (1964). Daily measurements of activity over a period of 10 days revealed variations in enzyme activity which are difficult to explain. However, the variations are smaller than those established in earlier tests. In order to obtain statistically valid results, the highest possible number of test fishes (at least 10 individuals for each measurement) must be investigated over a period of 5 to 10 days. NADP⁺-ICDH of individuals acclimated to 10°C showed 28% higher specific and non-specific activity than that of fish maintained at 20°C (experimental temperature 25°C). At 5 different adaptation temperatures, increasing activities were observed with decreasing adaptation temperature (compensation). These measurements were made over a period of 5 successive days.