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

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• 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.

     

Temperature acclimation in respiratory and cytochrome c oxidase activity in common carp (Cyprinus carpio)

• 1.1. Common carp (Cyprinus carpio) exposed to experimental temperatures of 12, 18, 24, 30 or 36°C for a 4-week period were used to investigate the effect of temperature acclimation on the frequency of opercular movement (FOM), growth and cytochrome c oxidase (CCO) activity in heart, liver and muscle.
• 2.2. An exponential relationship between FOM and temperature after the first week (10₁₀ =1.76) disappeared after the second week.
• 3.3. The initially high FOM at temperatures of 30 or 36°C and the low FOM at 18 or 12°C changed over 4 weeks to approach the FOM of fish at 24°C.
• 4.4. This change in the relationship of FOM to temperature from highly dependent to independent appeared to be thermal compensation.
• 5.5. Heart and liver CCO activities were significantly affected by temperature, with the lowest activity at the approximate optimum temperature for growth, 24°C.
• 6.6. Highest CCO activities for heart and liver occurred at both the highest and lowest temperatures.
• 7.7. Among the three tissues, heart CCO activity was generally the highest and most affected by acclimation temperature.
• 8.8. Muscle tissue had the lowest CCO activity and was unaffected by temperature.
• 9.9. The high CCO activity at a cold acclimation of temperature 12°C was probably due to thermal compensation and the high activity at 36°C may have been a result of thermal stress.

     

The effects of temperature and moisture on survival capacity,cuticular permeability,hemolymph osmoregulation and metabolism in the scorpion,Centruroides hentzi (banks) (scorpiones,buthidae)

• 1.1. The temperature and water relations of Centruroides hentzi females were investigated. At 12 and 72% relative humidity (RH), the lower and upper Lt₅₀ were -4.5 and 43.7°C, and -4.7 and 45.1°C, respectively. When exposed to high temperature stress, survivorship was significantly greater under mesic conditions.
• 2.2. Cuticular water loss was higher under xeric conditions (12% RH), ranging from 0.061 mg/cm²/hr at 30°C to 0.211 at 41°C.
• 3.3. Exposure to dry air (0–5% RH) resulted in a significant increase in hemolymph osmolality: from 441 to 688 mOsm over a 5 day period.
• 4.4. Mean oxygen consumption rates increased from 161.7 mm³/g/hr at 34°C to 541.6 at 44°C. ATPase activity was significantly higher in animals acclimated and tested at 35°C.

     

Changes in composition and proteolytic enzyme activities of artemia during early development

• 1.1. The proximate composition, total and free amino acids, and proteases of Artemia nauplii were determined during early development.
• 2.2. Moisture increased from 71.0% to 80.8%, crude protein decreased from 13.2% to 8.8%, crude fat and ash varied slightly.
• 3.3. The total amino acids decreased. Free amino acids changed in three patterns.
• 4.4. Trypsin, chymotrypsin, carboxypeptidase A, B and cathepsin B and C increased in activity. The activity of trypsin was lower, while cathepsin B and C were the highest.
• 5.5. The protease activities were maximal at pH 7.5 and 8.0, and at 45°C on casein.
• 6.6. The optimal pH for carboxypeptidase A was 4.0, for carboxypeptidase B was 4.5, for trypsin and chymotrypsin were 7.0–7.5. The protease(s) active at pH 9.0–9.5 were to be determined.

     

Purification and characterization of the amylolytic enzymes of Saccharomycopsis fibuligera

• 1.1. A complex of extracellular amylolytic enzymes produced by Saccharomycopsis fibuligera KZ, grown on fine fibre (waste product from corn starch production) and corn-steep liquor, has been studied.
• 2.2. α-Amylases and glucoamylases, as the main representatives of this complex, were separated by hydrophobic chromatography on Spheron 300 LC.
• 3.3. Individual isoenzymes of one type were separated on FPLC-Mono Q.
• 4.4. The relative molecular weight of α-amylases is 54,000, glucoamylases 62,000, maximal activity is reached by both enzymes between pH 5.0 and 6.2 at a temperature of 40–50°C.
• 5.5. Glucoamylases have a higher stability of the native structure than α-amylases, they retain 55% of their original activity, even after 10 min of incubation at 100°C.

     

Transfer system of cobalamin from pellicle to cytosolic binding proteins in Euglena gracilis

• 1.1. A cobalamin (Cb1)-transfer system from the pellicle to cytosolic Cb1-binding proteins occurs in Euglena gracilis.
• 2.2. The Cbl-transfer activity showed thermal dependency. The optimum temperature was 50°C. The Cbl-transfer activity was increased significantly above pH 7.0.
• 3.3. ATPase, thiol-groups and metal ions were not involved in the Cbl transfer.

     

The digestive enzymes of the pacific brown shrimp Penaeus californiensis.: I—Properties of amylase activity in the digestive tract

• 1.1. Crude extract of the whole digestive tract from the brown shrimp (P. californiensis) was investigated for digestive amylase activity.
• 2.2. Considerable amylase activity was found at pH 6.5–8.0, with optimum pH at around 7.5.
• 3.3. Optimum temperature was found between 30–40°C, similar to amylases from other crustaceans.
• 4.4. Amylase activity was highly halotolerant, having 50% maximum activity at 3 M NaCl.
• 5.5. Maximum amylase activity was found at 0.01 M NaCl.
• 6.6. Amylase activity was partially inhibited by the divalent ions Hg²⁺, Zn²⁺, Cu²⁺ and Cr²⁺.
• 7.7. Mg²⁺ and Ca²⁺ ions seemed to enhance amylase activity.

     

The effect of cortisol on stimulation of enzymatic activity and absorption of amino acids in the small intestine of adult hamsters

• 1.1. The s.c. administration of cortisol to hamsters (50 mg/kg body wt/day for 4 days) produces a significant increase in maltase sucrase, alkaline phosphatase and leucineaminopeptidase activity in intestinal mucosa.
• 2.2. Lactase activity is unaffected by cortisol.
• 3.3. Gamma-glutamyltranspeptidase activity increases slightly in females but remains unchanged in males.
• 4.4. Cortisol causes increase in proline and glycine absorption without changing the absorption of lysine.

     

Phenoloxidase activity of acridid grasshoppers from the subfamilies melanoplinae and oedipodinae

• 1.1. Phenoloxidase activity and wound melanization was studied in five species of grasshoppers representing the subfamilies Melanoplinae and Oedipodinae.
• 2.2. Most of the phenoloxidase activity was detected in the plasma fraction of grasshopper whole-body homogenates and supernatant fractions of the hemolymph. The species representing the Oedipodinae had 20–50% higher percentage of the total phenoloxidase activity associated with particulate matter from a whole-body homogenate when compared to the Melanoplinae.
• 3.3. Phenoloxidase activity could not be detected in sclerotized cuticle of adult grasshoppers.
• 4.4. The phenoloxidase existed as a zymogen which could be activated by chymotrypsin and inhibited by KCN and NaCN while EDTA showed no effect. It had optimum activity at 37°C and pH 7.3.
• 5.5. These findings are discussed in relation to wound repair and immune responses to infection in grasshopper species.

     

The regulation of glucose 6-phosphate dehydrogenase from Dicentrarchus labrax (bass) liver

• 1.1. Kinetic constant values of the reaction catalyzed by bass liver glucose 6-phosphate dehydrogenase show to be modified between 10 and 40°C.
• 2.2. The Arrhenius plot between 10 and 50°C shows two slopes with different activation energies.
• 3.3. These results suggest a regulation of this enzyme by environmental temperature.
• 4.4. Kinetics of ATP inhibition were examined between pH 6.2 and 7.8: patterns and K_i values obtained are affected by the pH variation.
• 5.5. NADH is an effective inhibitor of bass glucose 6-phosphate dehydrogenase but this enzyme does not show NAD-linked activity.
• 6.6. Kinetics of pyridoxal 5′-phosphate inhibition have indicated the presence of a lysine in the catalytic site for NADP⁺.

     

Effect of temperature on oxygen consumption and heart rate of the australian crayfish Cherax tenuimanus (Smith)

• 1.1. Oxygen consumption at 18°C was 60% of the rate at 22 and 26°C.
• 2.2. Critical points, where the rate of oxygen consumption changed, were defined at 22°C (2.89 mg DO) and 26°C (3.46 mg DO). Linear regressions were fitted showing that oxygen consumption declined significantly (81.5% ±4.5) below the critical point.
• 3.3. Oxygen consumption was proportional to weight. Allometric relationships resulted in variable temperature-related coefficients for respiratory dependence on weight, a reflection of the crayfish adaptation towards re-establishment of a new equilibrium state.
• 4.4. Heart beat rate was lower at 18°C, and highest at the acclimation temperature (22°C). Stress at 26°C was evident.

     

Behavioural thermoregulation and critical thermal limits of Macrobrachium acanthurus (Wiegman)

• (1)The preferred temperatures of Macrobrachium acanthurus were determined for prawns acclimated to 20°C, 23°C, 26°C, 29°C and 32°C, and the final preferendum estimate was (29.5°C).
• (2)The critical thermal minima (CTMin) and maxima (CTMax) were 11.0°C, 12.1°C, 13.0°C and 14.8°C, and 34.2°C, 35.0°C, 36.1°C and 39.8°C, respectively.
• (3)The zone of thermal tolerance assessed using the CTMin and CTMax boundaries was 644°C².
• (4)The acclimation response ratio was between 0.33 and 0.62.
• (5)To cultivate this species in the southeastern region of México it should be done in not <15°C (CTMin) during the winter and below 38°C in summer (CTMax).

     

Effects of temperature and metabolic inhibitors on contraction of anterior byssus retractor muscle of Mytilus

• 1.1. Anterior byssus retractor muscle of Mytilus (ABRM) was stimulated to contract by ACh (acetylcholine) and effects of temperature (5–30°C), FDNB (1-fluoro 2,4 dinitro-benzene) and IAA (iodoacetic acid) on tension response were examined.
• 2.2. Isometric tension was highest at the temperature range of 10–20°C and decreased at higher and lower temperature than that range.
• 3.3. The rate of tension decay after washing of ACh was accelerated by the increase of temperature.
• 4.4. Tension redevelopment after release of 1 % during contraction was much smaller at 5°C than at 20°C.
• 5.5. Tension development by ACh and the rate of tension decay after washing of ACh were remarkably decreased by the treatment of FDNB or IAA.
• 6.6. The above results were discussed from the viewpoint that energy metabolism might be related to catch.

     

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.

     

Utilization of body stores in embryonated ova and larvae of two coregonid species (Coregonus lavaretus L. and C. albula L.)

• 1.1. Developing eggs of whitefish (Coregonus lavaretus L.) and vendace (Coregonus albula L.) were kept at 1–2°C and some eggs taken gradually up to 8°C to provoke mass hatching of embryos.
• 2.2. Wet weight, dry matter and the contents of lipid, protein and ash were measured in fish during the course of experiment.
• 3.3. Dry matter content decreased gradually in whitefish eggs from 15.64 to 11.95% during 1 month at 1–2°C, whereas vendace eggs showed only a slight decrease from 16.27 to 15.53%.
• 4.4. In both species protein content decreased but lipid increased when approaching the natural time of hatching.
• 5.5. During delayed hatching at low water temperatures protein contributes to catabolism, whereas lipid content decreased only in the later phase of the experiment.
• 6.6. Larvae starved for 10 days after hatching lost increasing amounts of dry matter (from 26.1 to 50.3% of body weight) and protein (from 18.7 to 45.9% of body weight) as they remained longer in cold water as embryos.
• 7.7. A correspondence was found between assessment of metabolic utilization of body stores based on chemical analysis of fish body and previous work on oxygen consumption and nitrogen excretion.

     

Isohematoporphyrin-diesters and diethers

• 1.1. A series of diesters of isohematoporphyrin (isoHp), from dimethyl to dioctyl were prepared according to Rimington et al. (1989b). Their optical absorption, fluorescence spectra and high performance liquid chromatography (HPLC) retention times were recorded.
• 2.2. A plot of HPLC retention time against number of C atoms in the alcohol used for esterification was approximately linear at first then rising steeply from diamyl to diocyi ester, whether a gradient elution was used or only methanol: water, 95/5, at pH 7.5.
• 3.3. Preparation of the diethers of isoHp was much more difficult than that of the corresponding derivatives of hematoporphyrin (Hp). Several different methods were investigated, varying both times and temperatures.
• 4.4. These methods included reaction of isoHp or its demethyl ester with
  • 4.1.(i) a bromoalkane in presence of anhydrous K₂CO₃;
  • 4.2.(ii) reaction with bromoalkane and Ag₂O;
  • 4.3.(iii) reaction of brominated-isoHp, prepared by using thionylbromide, with the selected alcohol, or corresponding sodium alcoholate;
  • 4.4.(iv) heating of isoHp alone with an alcohol containing 20% (w/v) H₂SCO₄ (temp. range from 45° to 118°C),
  • 4.5.(v) refluxing as in (iv) at the b.p. of the alcohol; and
  • 4.6.(vi) carrying out this reaction in refluxing ethyleneglycoldimethyl ether (b.p. 85°C) or diethyleneglycoldimethyl ether (b.p. 155°C).
• 5.5. Some diether formation was observable by all these methods but yields were small, a considerable quantity of unreacted isoHp and other products remaining.
• 6.6. Examined by HPLC, the diethers consistently afforded a forked peak which on thin layer chromatography was only resolved into two very closely associated bands by a solvent mixture carefully selected for development.
• 7.7. On elution these materials had virtually identical optical absorption and fluoresence spectra.
• 8.8. The nature of the association is discussed, atropisomers (Gottwald and Ullman, 1969) and possible stacked monomer: dimers (Abraham et al., 1963) being considered as possibilities.

     

Activity of NAD-sorbitol dehydrogenase is caused by biosynthesis of enzyme protein in diapause and non-diapause eggs of the silkworm,Bombyx mori

• 1.1. Using SDS-PAGE and immunoblotting analyses with anti-sorbitol dehydeogenase (EC 1.1.1.14, SDH) serum, changes in amount of SDH protein were examined in diapause and non-diapause eggs of the silkworm, Bombyx mori.
• 2.2. When diapause eggs were exposed to 5°C from 2 days after oviposition to break the diapause gradually, SDH protein appeared after 50-day chilling, and then the amount increased along with chilling period. This changing pattern paralleled that in SDH activity.
• 3.3. In diapause eggs treated with HCl after chilling at 5°C for 30 days to break the diapause quickly, and non-diapause eggs, changing patterns in amount of SDH protein also paralleled those in SDH activity.
• 4.4. These results showed that SDH activity was caused by biosynthesis of SDH protein, independent of diapause or non-diapause eggs.
• 5.5. Occurrence of SDH correlates with the three developmental phases: diapause termination, embryonic growth, and larval differentiation. In larva, SDH was mainly localized in the fat-body.

     

Sea-water tolerance in freshwater-resident arctic charr (Salvelinus alpinus)

• 1.1. Freshwater-resident Arctic charr acclimated for 2 months at 8°C, 15% were divided into four experimental groups in July and exposed to 1 and 8°C in 15 and 34% salinity.
• 2.2. Only slight changes in gill Na-K-ATPase activity, blood plasma osmolality and blood plasma concentrations of Cl⁻ and Mg²⁺ were found for the fish exposed to 1 or 8°C in brackish water.
• 3.3. When exposed to sea-water at 8°C, an increase in osmolality and in concentrations of Cl⁻ and Mg²⁺ took place during the first 2–3 days, after which it levelled off.
• 4.4. If exposed to sea-water at 1°C, however, marked increases were found for all parameters measured and all the fish were dead within 5 days of exposure.
• 5.5. These results show that freshwater-resident Arctic charr—if acclimated to brackish water—can survive in sea-water during summer if the environmental temperature is not too low.

     

Influence of environmental factors on the deep and skin temperature in the European bison,Bison bonasus (L.)

• 1.1. In 43 European bison divided into three groups (Group A, 3–8-month-old calves; Group B, 18-month-7-year-old young bison; Group C, 12–24-year-old bison) the rectal, humerus region and abdomen region temperatures were measured.
• 2.2. The experiments were carried out in winter months, from mid-December to mid-March.
• 3.3. The mean rectal temperatures changed from 38.55°C in calves to 38.15°C in the oldest bison.
• 4.4. The mean temperatures of the humerus region changed from 20.69°C in calves to 21.49°C in older bison.
• 5.5. The mean temperatures of the abdomen region changed from 20.79°C in calves to 22.17°C in older bison (Gr. B).
• 6.6. The cluster analysis divided the bison into four groups named hot, warm, cool and cold bison.
• 7.7. Only air temperature measured 2 m above the ground and snow cover influenced the integrated bison temperature. Age, sex and mass as well as some environmental factors had no influence.
• 8.8. Measurements made 1 to nearly 4hr after a bison's death showed a drop in rectal temperature and mostly increases in temperatures of the humerus and abdomen regions.

     

Thermal properties for digestive enzymes of a sub-antarctic beetle,hydromedion sparsutum (coleoptera,perimylopidae) compared to those in two thermophilic insects

• 1.1. Proteolytic, lipolytic, amylolytic and cellulolytic activities were studied in adults of the phytophagous beetle, Hydromedion sparsutum, indigenous to the sub-Antarctic island of South Georgia.
• 2.2. Gastric enzyme activities were measured at experimental temperatures of 5–40°C and results were compared with those obtained from two thermophilic insects, Gryllus bimaculatus and Tenebrio molitor.
• 3.3. Protease and lipase activities in Hydromedion were 10–15 times lower than in Gryllus and Tenebrio.
• 4.4. In the temperature range of 5–15°C, α-amylase activity from Hydromedion was only slightly lower than that from Gryllus.
• 5.5. Hydromedion gut homogenates exhibited a distinct cellulolytic activity, even at a low temperature of 5°C.
• 6.6. Cellulolytic activity in the digestive tract of Hydromedion was confirmed by the evolution of ¹⁴CO₂ after consumption of labelled cellulose.
• 7.7. The thermal properties of digestive enzymes agree well with the role of Hydromedion as primary decomposer in its ecosystem.