The biochemical ecology of Biomphalaria glabrata,a freshwater pulmonate mollusc: The uptake and assimilation of exogenous glucose and maltose

• 1.1. The rates at which the pulmonate snail B. glabrata takes up soluble glucose and maltose from a defined medium were evaluated by measuring the buccal mass pulsation rate, the drinking rate, net changes in the concentrations of sugars in the medium and the rate of accumulation of ¹⁴C labelled maltose.
• 2.2. It was demonstrated that B. glabrata was capable of net accumulation of maltose and glucose via the mouth and integument, respectively.
• 3.3. Some of the maltose in the medium was also hydrolysed to glucose by exogenous snail enzymes.
• 4.4. The mechanisms involved in the accumulation of the sugars and the relevance of the results to the biochemical ecology of the snails and other aquatic invertebrates are discussed.

     

Characteristics of Crassostrea virginica crystalline style chitin digestion

• 1.1. Fundamental chitin digestion characteristics of Crassostrea virginica crystalline style were investigated.
• 2.2. Optimum temperature and pH were 34°C and 4.8. respectively.
• 3.3. The colloidal regenerated chitin (0.56mol/0.5 ml: GlcNAc equivalents) was saturating under all enzyme levels encountered.
• 4.4. There was no evidence of end product inhibition, even after 100 hr incubation.
• 5.5. Calculated K_m for the chitinase complex was 1.19mM when determined using a 30 min assay, but was only 0.70 mM when determined using a 4.6 hr assay.
• 6.6. Both K_m values are lower than reported for similar assays in other molluscs and for most bacteria.
• 7.7. Effect of substrate preparation on the kinetics are discussed.
• 8.8. Eight peaks of chitinase activity were resolved by DEAE-Fractogel ion exchange chromatography.

     

Lipoxygenase of the thermophilic bacteria thermoactinomyces vulgaris—properties and study on the active site

• 1.1. A lipoxygenase activity was purified from Thermoactinomyces vulgaris and some of its properties were characterized.
• 2.2. The enzyme showed a temperature activity range of 40–55°C with still significant activity over 60°C.
• 3.3. The pH of activity on linoleic acid had a broad range with an optimum at pH 6.0 and a weaker one at pH 11.0.
• 4.4. On arachidonic acid the pattern was narrow bell-shaped with an optimum at pH 6.5.
• 5.5. The purified lipoxygenase from Th. vulgaris showed an apparent K_m of 1 mM and V_max of 0.84 μmol diene/min/mg protein.
• 6.6. It was inhibited by the oxidation products, 9-HPOD and 13-HPOD.
• 7.7. A 160,000 Da molecular weight of the enzyme was determined by molecular filtration. Methionine, tyrosine, tryptophan and cysteine are apparently involved in its activity.

     

Laboratory studies on the oxygen consumption of the marine teleost,Lichia amia (Linnaeus, 1758)

• 1.1. The oxygen consumption of the marine teleost, Lichia amia was investigated under controlled laboratory conditions.
• 2.2. The routine oxygen consumption showed a strong circadian rhythm with the fish being mainly active during the light period.
• 3.3. The specific mass exponent (dimension: μg O₂/g/hr) is temperature independent and ranges from 0.27–0.29.
• 4.4. Starving the fish results in a mean decrease in active, routine and standard oxygen consumption of 21%, 24% and 20%, respectively.
• 5.5. Feecling led to an increase in the oxygen consumption of the teleosts, with the mean metabolic rate over the 24 hr that followed, being 58% and 50% higher for fish that had been starved for 162hr and 40 hr, respectively.
• 6.6. Apparent SDA showed some variation and ranged from 6.0 to 35.5%.
• 7.7. The results obtained are generally in agreement with those recorded for other teleosts.

     

The influence of aerial exposure on gas exchange and cardiac activity in the shore crab,Carcinus maenas (L.)

• 1.1. The cardiovascular physiology of adult Carcinus maenas (L.) emerging into air has been investigated at three different air temperatures.
• 2.2. Transition from seawater to air or vice versa triggered transient increases in cardiac and locomotor activity.
• 3.3. However, crabs became inactive 5–10 min after emerging from seawater (15°C) into air at the same temperature (15°C) or at lower temperatures (12–13°C) and heart rate fell.
• 4.4. At higher air temperatures (18–20°C) heart rate rose but to a lesser extent than predicted from aquatic Q₁₀ heart-rate values.
• 5.5. Crabs were again quiescent in aerial conditions.
• 6.6. Mean arterial oxygen tension (Pa_o2) was ~ 74 mmHg in submerged crabs but fell to ~ 38 mmHg in air while mean arterial carbon dioxide tension (Pa_o2) increased from 1 to 4 mmHg resulting in respiratory acidosis.
• 7.7. A model of gill function is proposed to explain the development of internal hypoxia in air.
• 8.8. The results are discussed in relation to the distribution of adult and juvenile C. maenas in situ.

     

The flow theory of enzyme kinetics: Role of solid geometry in the control of reaction velocity in live animals

• 1.1. When blood flows, membranes are bombarded with ions etc., whose entry creates an ATP demand proportional to flow rate. Also proportional to flow rate is ATP production from oxidation of substrates [S] from the same blood volume.
• 2.2. O₂ is limiting and reaction velocity at rest (metabolic rate) is determined by flow rate, F, but not by [S].
• 3.3. Since resting blood O₂ A-V difference is about 5 vol%, 11 circulated produces about 0.25 kcal in mammals, birds or warm reptiles.
• 4.4. Where O₂ is not limiting, as in most amino acid deaminations, V = K F[S] with K a constant unrelated to K_m.
• 5.5. At equal blood vol/kg, solid geometry dictates that the average cross-sectional area of major vessels/kg will be an inverse function of body mass. The smaller the animal, the shorter the vessels, the “thicker” the vessels/kg body wt, and at any one blood pressure, the higher the flow/kg/hr. If a man's major vessels were equal in cross-section/kg to those of a shrew, it would take 2241 of blood to fill them.
• 6.6. Growth decreases flow/kg (and therefore metabolic rate), by decreasing vessel cross-section/kg without changing blood pressure or linear velocity of flow.
• 7.7. Surface area/g, body wt to some power, average vessel length/kg, circulation time and average major vessel cross-sectional area are all related mathematically.

     

A lipid transfer particle in Musca domestica haemolymph

• 1.1. In Musca domestica haemolymph a lipid transfer particle (LTP) is present.
• 2.2. Musca domestica LTP is able to catalyze the transfer of lipids between different housefly lipophorin forms and also between lipophorins of Diptera and Lepidoptera.
• 3.3. The lipophorin of larval Dione juno (Lepidoptera) was purified and is composed of two apolipoproteins, apolipophorin I (M_{r = 209,000}) and apolipophorin II (M_{r = 85,000}) with a density of 1.124 g/ml.
• 4.4. The density of housefly lipophorin undergoes variations during the gonotrophic cycle.
• 5.5. The lipophorin density variation results suggest that when a high rate of lipid utilization occurs, the lipophorin has a higher density value.

     

Ventilatory rate in the butterfish (Pholis gunnellus) as a consequence of temperature and previous emersion

• 1.1. Observation of ventilation in immersed Pholis gunnellus showed a linear relationship between ventilatory rate and temperature between 8 and 20°C.
• 2.2. At 13°C and after 30 min emersion, ventilatory rate was initially lower than prior to emersion, providing evidence of adequate uptake of O₂ for standard metabolism during the emersion period.
• 3.3. This species has a laterally elongate body form with reduced scales and extensive mucus secretion.
• 4.4. During emersion, gaping behaviour probably exposes the gills and extensively vascularised oesophageal regions to air.
• 5.5. These are considered to be morphological and behavioural adaptations by P. gunnellus, to aerial respiration in the intertidal habitats occupied by this species.

     

Characterization of NaK ATPase from the gills of the freshwater prawn Macrobrachium rosenbergii (de Man)

• 1.1. The specific activity of Na-K ATPase was determined from the microsomal preparation of gills dissected from adult Macrobrachium rosenbergii.
• 2.2. Maximal ATPase activity was achieved at a substrate concentration of 0.5 mM ATP.
• 3.3. Optimal enzyme activity was obtained at pH of 7.5.
• 4.4. The Arrhenius plot of Na-K ATPase activity revealed a marked discontinuity at 30°C. “Mg” ATPase activity did not exhibit a marked discontinuity.
• 5.5. The E_a for Na-K ATPase and “Mg” ATPase was 14.6 kCal/mole and 9.31 kCal/mole respectively. Q₁₀ values for Na-K ATPase was 2.34 and for “Mg” ATPase 1.65.
• 6.6. ATPase activity and gill homogenate protein concentration exhibited a linear relationship up to 130 μg protein/ml.
• 7.7. Na-K ATPase activity was inhibited by 10⁻³ M ouabain. It was equally inhibited by the removal of K⁺ from the reaction medium.

     

Decarboxylation of uroporphyrinogen I and III in 2,3,7,8-tetrachlorodibenzo-p-dioxin induced porphyria in mice

• 1.1. The decarboxylation of uroporphyrinogens I and III by porphyrinogen carboxy-lyase (EC 4.1.1.37) in mouse liver supernatant was compared in relation to substrate concentrations.
• 2.2. In this species uroporphyrinogen III was the best substrate judging by the criteria of K_m/V_max (estimated for total porphyrinogens) and was converted into coproporphyrinogen faster than its series I isomer.
• 3.3. The difference between the two isomers was mainly due to the first decarboxylation.
• 4.4. This difference was confirmed by calculation of the Hill coefficient and of Lineweaver-Burk plot which suggested that isomer I induced negative cooperativity in the active centre of the enzyme.
• 5.5. After treatment with a porphyrogenic dose of TCDD (25 μg/kg/week for 9 weeks) differences between uroporphyrinogen I and III as substrate were maintained.
• 6.6. In addition treatment reduced V_max and K_m (estimated for total porphyrinogens) of liver porphyrinogen carboxy-lyase to about half control values for both isomers.
• 7.7. V_max was reduced mainly because of the formation of smaller amounts of all products of decarboxylation, and K_m because more heptaporphyrinogen was formed than coproporphyrinogen.
• 8.8. Values of the Hill coefficient and Lineweaver-Burk plots suggested TCDD induced altered substrate affinity for isomer III too.
• 9.9. Treatment with TCDD did not affect the decarboxylation of uroporphyrinogen III by RBC porphyrinogen carboxy-lyase, estimated from K_m and V_max for total porphyrinogens formed.

     

IDentification of a microsomal retinoic acid synthase as a microsomal cytochrome P-450-linked monooxygenase system

• 1.1. To characterize an enzyme which metabolizes retinal in liver microsomes, several properties of the enzymatic reaction from retinal to retinoic acid were investigated using rabbit liver microsomes.
• 2.2. The maximum pH of the reaction in the liver microsomes was 7.6.
• 3.3. The K_m and V_max values for all-trans, 9-cis and 13-cis-retinals were determined.
• 4.4. The reaction proceeded in the presence of NADPH and molecular oxygen.
• 5.5. The incorporation of one atom of molecular oxygen into retinal was confirmed by using oxygen-18, showing that the reaction comprised monooxygenation, not dehydrogenation.
• 6.6. The monooxygenase activity was inhibited by carbon monoxide, phenylisocyanide and antiNADPH-cytochrome P-450 reductase IgG, but not by anti-cytochrome b₅ IgG.
• 7.7. The enzymatic activity inhibited by carbon monoxide was photoreversibly restored by light of a wavelength of around 450 nm.
• 8.8. The retinal-induced spectra of liver microsomes with three isomeric retinals were type I spectra.
• 9.9. The microsomal monooxygenase activity induced by phenobarbital or ethanol were more effective than that by 3-methylcholanthrene, clotrimazole or β-naphthoflavone.
• 10.10. These results showed that the monooxygenase reaction from retinal to retinoic acid in liver microsomes is catalyzed by a cytochrome P-450-linked monooxygenase system.

     

A novel inhibitor of glycogen phosphorylase from crayfish hepatopancreas

• 1.1. A novel glycogen phosphorylase inhibitor was partially purified from crayfish hepatopancreas.
• 2.2. The inhibitor was found only in two species of crayfish examined, and not in lobster, fresh and salt water clams, mussels or cockroaches.
• 3.3. The inhibitor is a small protein (M_r = 23,000) which did not show proteolytic activity.
• 4.4. Preliminary kinetic analysis of the inhibitory mechanism indicated that it bound to both glycogen and the glycogen phosphorylase protein.
• 5.5. Inhibitor binding to glycogen resulted in a competitive inhibition pattern with respect to glycogen phosphorylase (inhibition constant of ca 10 μg/ml).
• 6.6. The inhibitor also bound glycogen phosphorylase directly with a binding coefficient of 100 μg/ml resulting in a partially non-competitive inhibition pattern with respect to phosphate.

     

NADPH/NADP ratio could regulate the glyoxylate cycle in Tetrahymena pyriformis

• 1.1. The glyoxylic acid cycle pathway could be regulated through the modulation of the isocitrate dehydrogenase-NADP activity. This enzyme is inhibited by NADPH.
• 2.2. The effect on the glyoxylate cycle flux of variations in the rate of the NADPH-consuming pathways has been studied.
• 3.3. Increase in the rate of NADPH-consuming activity by addition of H₂O₂ produces inhibition of the glyoxylate cycle and decrease in the NADPH/NADP ratio.
• 4.4. These results suggest that the glyoxylate flux in Tetrahymena could be modulated by regulation of NADP-dependent isocitrate dehydrogenase by the NADPH/NADP ratio.

     

Metabolic roles of carnitine expressed through the carnitine acetyltransferase system in Candida pintolopesii

• 1.1. The carnitine-responsive mutant yeast, Candida pintolopesii ATCC 26014 and the wild type strain (ATCC 22987) were used to investigate the role of carnitine and the carnitine acetyltransferase system.
• 2.2. [³H]l-Carnitine, supplied to the cells, was incorporated into acetylcamitine and [¹⁴C]pantothenate was incorporated into CoA and its derivatives.
• 3.3. Both bioautography and quantitative assays indicated that the relative amounts of CoA and acetylCoA were very different in the mutant and wild type cells.
• 4.4. The wild type yeast maintained an acetylCoA/CoA ratio of 0.33 ± 0.09 indicating that most of the CoA in the cell is in the free CoA form. Carnitine was not required to establish this ratio nor did its presence lower it further.
• 5.5. In contrast, the mutant cells contained a high acetylCoA/CoA ratio (12.8 ± 3.0).
• 6.6. In the mutant cells, carnitine lowered the ratio by decreasing the intracellular acetylCoA concentration and releasing free CoA.
• 7.7. These data indicated that wild type yeast possess an effective mechanism that is not related to the CAT system for regulating the acetylCoA/CoA ratio.
• 8.8. This mechanism appears to be lacking in the mutant. The CAT system decreased the acetylCoA/CoA ratio in the mutant cells but not to the value which is found in the wild type strain.
• 9.9. In both stains of Candida pintolopesii, in the presence of carnitine, an acetylcamitine pool can be created whose concentration exceeds that of acetylCoA.
• 10.10. The intracellular apparent equilibrium constant (K_app) for carnitine acetyltransferase for wild type Candida pintolopesii ATCC 22987 was 0.73 ± 0.12, close to the established value of 0.6, indicating that the CAT system ran close to equilibrium.
• 11.11. The K_app for the CAT system of the carnitine-responsive mutant yeast was 7.7 ± 1.7 indicating that this reaction was not at equilibrium.

     

The sugar chemoreception niches of Bulinus (physopsis) globosus (morelet) and Bulinus rohlfsi (clessin), the two most important snail hosts of Schistosoma haematobium (Bilharz) in West Africa

• 1.1.The behavioural responses of two species of freshwater pulmonate snails Bulinus (P.) globosus and Bulinus rohlfsi] to sugar gradients were investigated by means of diffusion olfactometers.
• 2.2.Both snail species proved to be very discriminating in their responses. Of the 17 sugars tested, 35.3%, namely d(−)glucuronic, maltotriose, maltose, cellobiose, d(−)arabinose, d(+)mannose proved to be statistically significant attractants or arrestants to B. rohlfsi. Only 23.5% of these sugars (maltotriose, maltose), d(+) mannose and d(+) xylose were significant attraetants or arrestants to B. (P.) globosus.
• 3.3.Glucuronic acid was a significant repellent to B. rohlfsi but none of the sugars was a repellent to B. (P.) globosus.
• 4.4.The results are compared with those obtained for other snail species and their relevance to the ecology and control of the snails are discussed.

     

Hyaluronidase degradation of hyaluronic acid from different sources: Influence of the hydrolysis conditions on the production and the relative proportions of tetra- and hexasaccharide produced

• 1.1. Hyaluronic acid (HA) can be digested with a Streptomyces hyaluronidase.
• 2.2. The rate of production and the ratio of tetrasaccharide (T) and hexasaccharide (H), studied by HPLC, varied with the temperature and duration of hydrolysis.
• 3.3. The rates of production and the respective amounts of the two oligosaccharides depended on the rheological properties of the HA from different sources.
• 4.4. A close relationship was found between the initial rate of hydrolysis and the intrinsic viscosity of the HA (η_i).
• 5.5. Our data suggest that enzymatic degradation at a given pH value, temperature, and duration of hydrolysis is dependent on the conformation of HA.
• 6.6. Moreover, under given conditions, the relative proportions of the two oligosaccharides depend on the η_i and may also reflect the degree of hydrolysis of the substrate.

     

Purification and properties of tetralysine endopeptidase from Escherichia coli AJ005

• 1.1. A new tetralysine endopeptidase from Escherichia coli AJ005 has been purified about 135-fold.
• 2.2. The peptidase seems to be specific to tetralysine among lysine homopolymers.
• 3.3. The optimal pH was about 7.5
• 4.4. The activity was inhibited by KCN but not inhibited by soybean trypsin inhibitor.
• 5.5. The apparent K_m value was 2.5 × 1O⁻³ M for tetralysine.

     

A simple and reliable method for the purification of Pseudomonas aeruginosa phospholipase C produced in a high phosphate medium containing choline

• 1.1. Pseudomonas aeruginosa phospholipase C from culture supernatants of bacteria grown in high-P_i basal salt medium with choline, as the sole carbon and nitrogen source, was purified by precipitation with 70% saturation ammonium sulfate in the presence of celite.
• 2.2. The PLC activity was eluted of this mixture by the use of a reverse gradient of 70-0% ammonium sulfate.
• 3.3. The peak containing the PLC activity revealed a single protein after SDS-PAGE.
• 4.4. The method could also be applied to purify PLC produced in a low-P_i complex medium. The resultant preparation was not homogeneous.
• 5.5. The molecular weight for both PLC preparations was about 70 kDa.
• 6.6. Both PLC used phosphatydilcholine and sphingomyelin as substrates, displayed hemolytic activity an exhibited an apparent K_M of 25 mM for p-nitrophenylphosphorylcholine.
• 7.7. They were not inhibited by 1% sodium deoxycholate but were 30% inhibited by 1% Triton X-100.
• 8.8. 2% sodium dodecylsulfate and 1% tetradecyltrimethylammonium bromide inhibited the PLC from the HP_l-BSM plus choline but not the enzyme from the LP_l-CM.

     

Human GMP synthetase

• 1.1. The specific activity of GMP synthetase was measured in several human tissues and found to be highest in cultured skin fibroblasts, followed by bone marrow, leukocytes, erythrocytes. placenta, and liver.
• 2.2. The enzyme from fibroblasts was purified approximately 50-fold by ammonium sulfate fractionation and gel filtration.
• 3.3. The K_m values were determined to be 4.9μM for XMP, 270μM for ATP. and 340 μM for glutamine.
• 4.4. Ammonium sulfate could replace glutamine as the amino donor but was much less efficient.
• 5.5. The enzyme was specific for ATP as the energy source.
• 6.6. Unlike the calf thymus enzyme, the human enzyme has no requirement for a reduced sulfhydryl compound.
• 7.7. Human GMP synthetase is inhibited by ATP, dATP, azaserine, and hydroxylamine.

     

Factors affecting oxygen consumption in wild-caught yellow-bellied marmots (Marmota flaviventris)

• 1.1. All age groups gained mass during the active season, but mass-gain of adult females was delayed during lactation.
• 2.2. The relationship of body mass to metabolic rate varied widely; when the relationship was significant, R² varied from 10.3 to 72.6%. Body mass affects V_O2 more during lactation than at any other period.
• 3.3. Mean VinO₂ of adult males was higher in June than that of adult, non-lactating females.
• 4.4. V_O2 of reproductive females was significantly higher during lactation than during gestation or postlactation because specific V_O2 varied. Specific V_O2 of non-reproductive females declined over the active season.
• 5.5. Specific V_O2 of all age groups declined between the premolt and postmolt periods. The reduced maintenance costs can contribute 20–46% to daily growth.
• 6.6. Observed V_O2 was lower than the value predicted from intraspecific or interspecific Bm:M regressions.
• 7.7. V_O2 of wild-caught marmots was lower than that of marmots maintained in the laboratory, probably because of dietary differences.
• 8.8. Because basal metabolism is a stage on a food-deprivation curve, we suggest that basal metabolic rate is not an appropriate measure of the metabolic activity of free-ranging animals.