Effect of triton X-100 treatment on the lipid composition,biochemical and functional properties of hake (Merluccius hubbsi) myofibrils purified from pre- and post-spawned fish

• 1.1. The effects of Triton X-100 treatment on the lipid contents and functional properties of hake myofibrils from pre- and post-spawned fish were investigated.
• 2.2. Differences in lipids, biochemical and functional properties of hake myofibrils related to the gonadal condition of fish were observed.
• 3.3. Triton X-100 treatment removed 65% of polar lipids in myofibrils from pre-spawned fish and only 10% in myofibrils from post-spawned fish.
• 4.4. Triton X-100 increased the Hill coefficient to 1.5 in an allosteric type of reaction for the myofibrillar Mg²⁺-ATPase from pre-spawned hake.
• 5.5. The detergent effect observed on the contraction response was greater in myofibrils from prespawned fish than in post-spawned fish.

     

Subcellular distribution of metals within the kidney of the bivalve Mercenaria mercenaria (L.)

• 1.1. The subcellular distribution of nine transition metals (plus four additional elements) was measured in the kidney tissue of the quahog, Mercenaria mercenaria.
• 2.2. Elemental analyses of the subcellular fractions indicated three main patterns of metal distribution within kidney cells.
• 3.3. Barium, iron, manganese and lead were associated primarily with kidney granules.
• 4.4. Cadmium, copper, potassium and magnesium were found mainly in the cytosolic fraction.
• 5.5. Calcium, phosphorus and zinc were found in all isolated fractions, probably reflecting the important roles that these elements play in bivalve metabolism.
• 6.6. The organelle composition of the isolated subcellular fractions was determined using marker enzyme assays and microscopic techniques.

     

Partial purification and characterization of cysteine proteinase from metamorphosing tadpole tails of Rana catesbeiana

• 1.1. A leupeptin-sensitive proteinase was partially purified from regressing tadpole tails by acetone factionation and column chromatography on S-Sepharose.
• 2.2. The enzyme degraded hemoglobin and myoglobin at pH 3.0. The enzyme also hydrolyzed Z-Phe-Arg-MCA and Boc-Val-Leu-Lys-MCA at pH 4.0.
• 3.3. The enzyme activity was inhibited by leupeptin, egg cystatin, E-64 and monoiodoacetic acid and was activated by l-cysteine.
• 4.4. The enzyme degraded myosin and actin in myofibrils of tadpole tails.
• 5.5. The enzyme belongs to the cysteine proteinase and is possibly involved in tail degradation during the metamorphosis of tadpoles.

     

Carbonic anhydrase III content in various equine muscles

• 1.1. In this study, carbonic anhydrase III (CA-III) content in 18 equine muscles was determined by enzyme immunoassay.
• 2.2. It was found to differ in several muscles.
• 3.3. That in external intercostal muscle, rectus abdominis muscle and splenius muscle from four horses was very high.
• 4.4. Although the masseter muscle had only type I fibers, CA-III content was similar to that in mixed-fiber type muscles such as the biceps femoris muscle.
• 5.5. It thus appear that equine type I fibers can be further subgrouped.

     

Biomines-stimulated phosphorylation and (Na+, K+-ATPase in the gills of the chinese crab,Eriocheir sinensis

• 1.1. Subcellular distribution of (NA⁺, K⁺-ATPase and ouabain-insensitive ATPase (Mg²⁺-ATPase) are compared in branchial tissues of the euryhaline crab, Eriocheir sinensis, acclimated to fresh water.
• 2.2. Both the anterior and posterior gills contain cAMP-dependent protein kinase and endogenous protein substrate for phosphorylation.
• 3.3. Phosphorylation occurs in both “particulate” and “soluble” subcellular fractions but its stimulation by cAMP is restricted to the “soluble” fraction.
• 4.4. serotonin (5-HT) and dopamine receptors are present only in the “light particulate” fraction isolated from the posterior gills.

• 1.(a) Serotonin and dopamine have no effect on the phosphorylation observed in a subcellular fraction alone.
• 2.(b) Activation of the phosphorylation by serotonin and dopamine is found when the soluble fraction (source of cAMP-dependent protein kinase) is added to the fraction P₃ from the posterior gills.
• 3.(c) No activation occurs with the fractions P₃ as well as P₁ or P₂ (not shown) from anterior gills of fresh water crab.
• 4.(d) Cyproheptadine, a serotonin receptor antagonist, inhibits the 5-HT dependent increase in phosphorylation.
• 5.(e) The dopamine receptor antagonist, chlorpromazine, inhibits dopamine-stimulated phosphorylation.
• 6.5. Ouabain mimics the effect of cyproheptadine on the serotonin-stimulated phosphorylation found in the posterior gills.

     

Proteolysis of skeletal muscle in yellowfin tuna (Thunnus albacares): Evidence of calpain activation

• 1.1. White muscle of yellowfin tuna is subject to a form of deterioration known as “burnt tuna”.
• 2.2. TEM and SDS-PAGE were used to quantify cellular differences in deteriorated white muscle of yellowfin tuna.
• 3.3. Electron micrographs showed a significant loss of Z-disc integrity and an increase in intracellular edema in burnt tuna.
• 4.4. Electrophoresis established that a specific doublet of proteins, 42 kD and 46 kD was lost.
• 5.5. Proteolysis of isolated myofibrils incubated in calpain (EC 3.4.22.17) was greatest at pH 7.5 and was selective for intermediate molecular weight proteins.
• 6.6. This evidence suggests that burnt tuna is a specific and limited proteolysis of myofibrillar structural proteins characteristic of calpain proteolysis.

     

Localization of the glucocorticoid receptor in rat liver cells: Evidence for plasma membrane bound receptor

• 1.1. The cytoplasmic glucocorticoid receptor of rat liver cells is in part recovered in the plasma membrane fraction.
• 2.2. After in vivo administration of [³H]dexamethasone, 0.35% of the radioactivity recovered is bound on plasma membranes.
• 3.3. Dexamethasone also binds in vitro specifically to plasma membranes. Expressed as fmol/mg protein, binding of dexamethasone to plasma membranes is comparable to binding to the soluble cytoplasmic fraction (cytosol).
• 4.4. Using polyclonal antibody to the glucocorticoid receptor and the indirect immunofluorescence technic, an intense decoration of the plasma membranes is observed, denoting a high concentration of glucocorticoid receptor on plasma membranes.
• 5.5. The localization of the receptor on plasma membranes could be of potential importance for its interaction with agents (mitogens, growth factors) initially acting on the cell membrane, regulating subsequent cell proliferation and growth at the level of the cell nucleus.

     

Characterization of Colorado potato beetle lipophorin: A hydrocarbon-rich diacylgycerol-poor lipophorin

• 1.1. The lipid composition of lipophorin from the Colorado potato beetle, Leptinotarsa decemlineata Say, was analyzed.
• 2.2. This insect lipophorin contains 44% lipid and is characterized by large amounts of hydrocarbons and small amounts of diacylglycerol.
• 3.3. This is the first observation of a diacylglycerol-poor insect lipophorin in haemolymph.
• 4.4. Since the main energy source for flight in the Colorado potato beetle is proline, the low diacylglycerol content in lipophorin must be related to its peculiar flight metabolism.
• 5.5. This lipophorin, however, can still take up appreciable amounts of diacylglycerol from the locust fat body. Hydrocarbon uptake by this lipophorin was also demonstrated.
• 6.6. The main function of this lipophorin therefore seems to be transport of hydrocarbons from oenocytes to the cuticle.

     

Morphological and physiological adaptations in the marginal sphincter of anemone Phymactis clematis,dana 1849 from different environments

• 1.1. We studied the morphology and contractile properties in marginal sphincters isolated from anemones from different environments.
• 2.2. Sphincters from specimens from protected areas are ovoid or roughly square shape. Oval sphincters have increased number of mesogloeal branches and the main axis is thickened. Roughly square sphincters have irregular borders, mesogloeal axes of uniform thickness and homogeneous branching.
• 3.3. Specimens from exposed areas have sphincters with an ovoid shape and dichotomous branching.
• 4.4. Sphincters of specimens from partially protected areas show transition forms.
• 5.5. Under stimulation with KCl at different concentrations, sphincters of anemones from exposed environments contract faster and develop higher isometric forces than muscles isolated from specimens of protected areas.
• 6.6. It was concluded that sphincters of anemones from different environments have a morphology and a physiological response adapted to the milieu.

     

Thermodependence of the tyrosine transaminating activity in mitochondria of Drosophila Melanogaster

• 1.1. The effect of temperature on activation energy of enzyme was examined.
• 2.2. Irrespective of the method applied for solubilization of mitochondria or of insect developmental stage, a biphasic Arrhenius plot with a well defined break point was obtained.
• 3.3. Both the temperature at which the break point was observed and the activation energy above and below this point were dependent on the detergent used and the stage of insect development.

     

Hydrolysis of glycol chitin by chitinolytic enzymes

• 1.1. The hydrolysis of glycol chitin preparations by several β-N-acetylglucosaminidases was monitored colorimetrically with the potassium ferriferrocyanide reagent.
• 2.2. Glycol chitin samples from crab and insect sources varied considerably in chemical composition and susceptibility to enzymatic hydrolysis.
• 3.3. Insect endochitinase preferred crab glycol chitin as substrate while hen's egg white lysozyme preferred commercial glycol chitin.
• 4.4. Insect glycol chitin was well hydrolyzed by both enzymes.
• 5.5. Insect exochitinase did not digest glycol chitin.

     

Domain structure of the 90-kDa stress protein: Heparin- and antibody-binding domain

• 1.1. To understand the physiological roles of the 90-kDa stress protein (HSP90), we investigated the heparin- and antibody-binding domains of the protein.
• 2.2. For heparin-binding sites, HSP90 was digested completely with trypsin, and the digests were applied to a heparin-Sepharose column and eluted with 1.0 M NaCl, followed by 8.0 M urea.
• 3.3. Each elutant was purified by a reverse-phase C₁₈ column.
• 4.4. Two peptides from the NaCl-eluted fraction and no peptide from the urea-eluted fraction were purified.
• 5.5. The purified peptides were sequenced by an automated peptide sequencer.
• 6.6. One of the heparin-binding sites was present between Leu-362 and Arg-365; another was present between Leu-645 and Lys-648.
• 7.7. These two peptides were basic and considerably hydrophilic.
• 8.8. For antibody-binding sites, HSP90 was mildly digested with trypsin, electrophoresed on SDS-polyacrylamide gels and transferred to PVDF membranes.
• 9.9. The four bound of the trypsin fragments could be sequenced with a peptide sequencer.
• 10.10. There was only one antibody-binding peptide, 38 kDa, starting from Pro-2. The others showed no cross-reactivity with the antibody and started from Leu-283.
• 11.11. Therefore, the epitopes of HSP90 are present between Pro-2 and Leu-282.
• 12.12. The heparin-binding sites are present from the middle region of the HSP90 molecule, and the antigen sites are at the N-terminal domain.

     

Haematology of the australian eastern quoll,Dasyurus viverrinus

• 1.1. A variety of haematological parameters were determined in adult Dasyurus viverrinus.
• 2.2. Haemoglobin and red cell counts were high with a very low mean cell volume.
• 3.3. Basophils are absent but the eosinophils contain small numbers of basophilic granules which may indicate a dual role for this cell.
• 4.4. “Ring Form” leucocytes are present.
• 5.5. Three types of red cell picture could be identified, some animals showing large numbers of spherocytes, spicule cells, and inclusion bodies.
• 6.6. These cells resemble those found in some inherited human haemolytic anaemias but there was no evidence of haemolysis in the animals.
• 7.7. An alkali resistant haemoglobin component is present.

     

Induction kinetics of immune antibacterial proteins in pupae of Galleria mellonella and Pieris brassicae

• 1.1. Pupae of Galleria mellonella and Pieris brassicae given an injection with live, non-pathogenic Enterobacter cloacae or abiotic foreign molecules induce an acquired immunity that corresponds with the synthesis of haemolymph proteins of antibacterial activity.
• 2.2. This humoral defensive response which persists for several days, differs quantitatively between insect species and between the inducers used, although very different foreign bodies induced the same immune proteins in both lepidopteran insects.
• 3.3. A stronger and longer lasting response was consistently noticed in pupae immunized with non-pathogenic bacterium than after sterile nutrient broth injections.
• 4.4. A demonstrably elevated activity of haemolymph lysozyme and trace activity of cecropins found in pupae of Galleria treated with saline W, a salt solution physiological to moths, disappear soon after 36 hr from injection.
• 5.5. In P. brassicae, however, sterile insect Ringer can give a varying, if present at all, immune response.
• 6.6. A mechanical injury (sterile wounding of insect body) can occasionally induce a similar but much weaker response.
• 7.7. The antibacterial activity was drastically reduced in Pieris or completely depressed in most pupae of Galleria when actinomycin D or cycloheximide was given at an early time post-immunization with E. cloacae.
• 8.8. It is concluded that the de novo synthesis of ribonucleic acid and immune proteins is required for expression of antibacterial activity in pupal haemolymphs.
• 9.9. The synthesis of an immune mRNA was completed about 7 hr after the injection of the immunizing bacteria.

     

Cellulose digestion by the firebrat Thermobia domestica

• 1.1. A highly efficient cellulose digestion could be demonstrated in a primitive insect species, Thermobia domestica (Thysanura:Lepismatidae), by the application of a uniformly ¹⁴C-labelled substrate.
• 2.2. Gut extracts exhibit distinct hydrolytic activities toward different cellulosic substrates (cellobiose, sodium carboxymethylcellulose and microcrystalline cellulose). Therefore, the complete cellular complex must be present.
• 3.3. Besides cellulases, several other carbohydrates occur in the digestive juice, thus reflecting the omnivorous feeding habits of the insect.
• 4.4. The crop was found to be the main site of carbohydrate digestiopn, also including cellulolysis.
• 5.5. It is very likely that the cellulolytic enzymes derive from the gut tissues of the firebrat.

     

Palmitic acid metabolism in hepatopancreas of the freshwater shrimp Macrobrachium borellii

• 1.1. The palmitic acid fate as substrate for the synthesis of either glycerides or other fatty acids was studied in vivo and in the microsomal fraction from hepatopancreas of Macrobrachium borellii.
• 2.2. Most of the palmitic acid administered in vivo circulated to the hepatopancreas, being incorporated mainly in the triacylglycerol (TG) fraction.
• 3.3. Palmitic acid transformations into palmitoleic, stearic and oleic acids were observed in the hepatopancreas.
• 4.4. The in vitro biosynthesis of TG in hepatopancreas was more active than in other tissues. In the microsomal fraction, palmitic acid was also incorporated mainly in TG, and followed the α-glycerophosphate pathway.

     

The slow muscles in the uropod of crayfish,Procambarus clarkII

• 1.1. The fiber types of uropod muscles of the crayfish, Procambarus clarkii, were determined by the myofibrillar ATP-ase histochemistry and electrophysiology.
• 2.2. The ATP-ase histochemistry was carried out on the sections of the whole tailfan and the slow muscles were identified as being of the slow type on the basis of their low staining intensities.
• 3.3. The location of slow bundles in the mixed type muscles i.e. the dorsal rotator (DRT), the ventral rotator (VRT), and the telson-uropodalis posterior (TUP) was confirmed.
• 4.4. TUP was newly revealed in this study to be a mixed muscle.

     

2-Deoxyglucose transport by the frog sartorius: Effects of electrical stimulation and N-carbobenzoxy-glycyl-l-phenylalaninamide

• 1.1. Studies characterizing glucose transport in the frog sartorius were performed.
• 2.2. For nonstimulated and stimulated muscles, intracellular 2-deoxyglucose exceeded 2-deoxyglucose-6-phosphate at 15 min, showed little further increase, and was maintained below the extracellular concentration for 2 hr.
• 3.3. Accumulated 2-deoxyglucose-6-phosphate did not inhibit glucose transport.
• 4.4. Unlike in adipocytes, basal and stimulated 2-deoxyglucose transport showed no difference in sensitivity to N-carbobenzoxy-glycyl-l-phenylalaninamide.
• 5.5. Phenylarsine oxide blocked contraction-enhanced 2-deoxyglucose uptake.
• 6.6. These results suggest that the glucose transporter of the sartorius exhibits auto-regulation, and that basal transport is not regulated by the same process as in adipocytes.

     

Metabolic and contractile differentiation of rabbit muscles during growth

• 1.1. A study was carried out of post-natal evolution of the oxidative, glycolytic and contractile capacities in various types of rabbit muscle.
• 2.2. At birth, muscles are non-differentiated and present very limited metabolic and contractile activity, metabolism is mainly oxidative in all muscles.
• 3.3. Although muscular discrimination is manifest from the sixth week after birth, the glycolytic metabolism reaches its maximum capacity only after six to eight weeks.
• 4.4. Subsequently, oxidative metabolic capacity steadily decreases until adulthood.

     

Characterization of two distinct latent proteinases associated with myofibrils of crucian carp (Carassius auratus cuvieri)

:

• 1.1. Enzymatic properties of two distinct proteinases tightly associated with crucian carp myofibrils were characterized.
• 2.2. These proteinases were latent but activated at 50 and 60°C, respectively.
• 3.3. The optimum pH of 50°C-proteinase was neutral-alkaline, while that of 60°C-proteinase was weak acid-neutral pH.
• 4.4. Both proteinases required more than 1% NaCl for the activity, but 50°C-proteinase was partially inhibited at higher concentrations of NaCl.
• 5.5. Both proteinases were regarded as trypsin-like proteinases belonging to a serine proteinase family, but only 60°C-proteinase was sensitive to urea, n-butanol and iso-propanol.