Comparative nuclear magnetic resonance studies of diffusional water permeability of red blood cells from different species. V—Rabbit (Oryctolagus Cuniculus)

• 1.1. The diffusional water permeability (P_d) of rabbit red blood cell (RBC) membrane has been monitored by a doping nuclear magnetic resonance (NMR) technique on control cells and following inhibition with p-chloromercuribenzene sulfonate (PCMBS).
• 2.2. The values of P_d were around 6.3 × 10⁻³ cm/sec at 15°C, 7.0 × 10⁻³cm/sec at 20°C, 8.0 × 10⁻³ cm/sec at 25°C, 9.1 × 10⁻³ cm/sec at 30°C and10.7 × 10⁻³ cm/sec at 37°C.
• 3.3. Systematic studies on the effects of PCMBS on water diffusion indicated that the maximal inhibition was reached in 15 min at 37°C with 0.5 mM PCMBS.
• 4.4. The values of maximal inhibition were around 71–74% at all temperatures.
• 5.5. The basal permeability to water was estimated as 1.6 × 10⁻³cm/sec at 15°C, 2.0 × 10⁻³cm/sec at 20°C, 2.4 × 10⁻³cm/sec at 25°C, 2.6 × 10⁻³cm/sec at 30°C, and 3.1× 10^{−3 cm/secat 37°C}.
• 6.6. The activation energy of water diffusion was around 18 kJ/mol and increased to 27 kcal/mol after incubation with PCMBS in conditions of maximal inhibition of water diffusion.
• 7.7. The membrane polypeptide electrophoretic pattern of rabbit RBCs has been compared with its human counterpart.
• 8.8. The rabbit membrane contained a higher amount of spectrin (bands 1 and 2), while the band 6 (glyceraldehyde-3-phosphate dehydrogenase) was markedly less intense.
• 9.9. Considerable differences in the electrophoretic patterns of the two sources of RBC membranes appeared in the bands migrating in the band 4.5 region and in front of band 7, where some polypeptides were apparent in higher amounts in the rabbit RBC membrane.

     

Soluble cyclic amp-dependent protein kinases from chick liver: Effects of NaCl and heat-stable modulator

• 1.1. Two cyclic AMP-dependent protein kinases—Fraction I and II—have been isolated from chick liver soluble preparation on DEAE-cellulose.
• 2.2. Both fractions have an apparent K_m for ATP of 2 × 10⁻⁶M, are stimulated maximally by 5 × 10⁻⁸ M cyclic AMP and phosphorylate mainly basic proteins—histone and protamine.
• 3.3. They exhibit various pH values for optimal activity and show differences with respect to both sensitivity to NaCl and substrate specificity.
• 4.4. The heat-stable protein modulator inhibits the cyclic AMP-dependent protein kinase activity of both fractions, but with cyclic GMP one kinase is stimulated and the other inhibited.
• 5.5. Slight differences in histone triggered holoenzyme dissociation as well as the lack of difference between their ability for subunit reassociation do not allow to classify these isozymes as protein kinases of Type I and II, according to Corbin et al. (1975).

     

On the role of fumarate reductase in anaerobic carbohydrate catabolism of Mytilus edulis L

• 1.1. The role of the fumarate:NADH oxidoreduction in the anaerobic glycolysis of the sea mussel is examined and discussed.
• 2.2. Fumarate reductase activity is present in submitochondrial particles especially from adductor muscle, digestive gland and mantle.
• 3.3. The pH optimum of the enzyme complex is 7.9; the approx K_m's for NADH and fumarate are 4.0 × 10⁻⁵ M and 6.3 × 10⁻⁵ M, respectively.
• 4.4. The enzyme complex is inhibitied by amytal, antimycin, ethanol, malonate, phosphate, rotenone, and succinate, and stimulated by Mg²⁺.
• 5.5. It is concluded that part of the mitochondrial respiratory chain is involved in the reduction of fumarate by NADH, comprising site 1 of the oxidative phosphorylation.

     

Intracellular localization and characterization of 3-hydroxykynureninase in human liver

• 1.1. 3-hydroxykynureninase in human liver was present in cytosol and mitoehondria.
• 2.2. The cytosolic enzyme and mitochondrial enzyme had the same physiological and enzymic properties.
• 3.3. The enzyme had a mol. wt of 130,000 by gel filtration and isoelectric point of pH 5.9.
• 4.4. The enzyme was active for 3-hydroxykynurenine and kynurenine, and its activity ratio was 15:1. The apparent K_m values of the enzyme were 7.7 × 10⁻⁵M for 3-hydroxykynurenine, 1.0×10⁻³M for kynurenine and 2.5 × 10⁻⁶M for pyridoxal 5'-phosphate with 3-hydroxykynurenine.
• 5.5. Some other properties of purified enzymes are described.

     

The effect of temperature on the acid-base status of the blood of the hatching quail (Coturnix c. japonica)

• 1.1. The ambient temperature of embryos of pipped eggs was reduced from 38 to 28°C for a period of 45 min.
• 2.2. The blood P_CO2 was lower and the blood more alkaline at 28°C than at 38°C.
• 3.3. At 28°C plasma [HCO₃⁻] ] was lower than predicted from the blood buffer line determined in vitro.
• 4.4. The plasma concentrations of strong ions and lactate were the same at both temperatures.
• 5.5. After the ambient temperature had been returned to 38°C for a period of 45 min, blood pH was more acidic than before cooling, but there was no difference in blood P_CO2.
• 6.6. The plasma [HCO₃⁻] was the same as that at 28°C and plasma [K⁺] was higher than before cooling.
• 7.7. The results arc discussed in relation to the factors affecting blood pH in embryos at this stage of development.

     

Respiration and energetics in west indian gorgonacea (Anthozoa,octocorallia)

• 1.1. The rates of oxygen consumption of five species of Gorgonacea were determined and their daily energy requirements for metabolism were estimated.
• 2.2. Oxygen consumption rates varied between 0.15 and 0.76 mg O₂ g organic matter⁻¹ hr⁻¹.
• 3.3. Daily energy requirements varied between 13 and 66 cal g organic matter⁻¹ d⁻¹.
• 4.4. Energy costs for maintenance were somewhat lower than in other reef-dwelling Anthozoa.

     

Respiration in the eastern water dragon,Physignathus lesueurii (agamidae)

• 1.1. Some aspects of the gas exchange system of a diving lizard, Physignathus lesuewii were studied.
• 2.2. Breathing patterns were analysed.
• 3.3. Breathing rate increases logarithmically with temperature and Q₁₀ = 1.8. LogBR = −0.237 + 0.0256 T.
• 4.4. Gas tensions in lung air and arterial and venous blood were measured. Arterial pH declines with increasing temperature.
• 5.5. Temperature has a marked effect on oxygen affinity of the blood (ΔH = −10.1 kcal mol⁻). A Bohr effect was also noted.
• 6.6. CO₂ equilibrium curves were drawn.
• 7.7. The results are considered with a view to anticipating the efficiency of the gas exchange system of this species under conditions of variable temperature and during diving.

     

Isolation and characterization of lipoamide dehydrogenase from mackerel dark muscle

• 1.1. Lipoamide dehydrogenase was purified 1500-fold from mackerel dark muscle.
• 2.2. The enzyme was homogeneous as judged by acrylamide gel electrophoresis in the presence and absence of SDS.
• 3.3. Molecular weights of 102,000 and 55,000 were estimated for the native and denatured enzyme, respectively.
• 4.4. Optimal activity for the enzyme was obtained at around pH 5.7 and enhanced with citri acid.
• 5.5. Loss of activity was less than 5% by incubating the enzyme at 70°C for 20 min.
• 6.6. An apparent K_m of 3.1 × 10⁻³ M was obtained for dl-lipoic acid and 1.5 × 10⁻⁵ M for NADH.
• 7.7. The properties of lipoamide dehydrogenase from mackerel dark muscle observed in this investigation were very similar to those reported for the enzyme from other sources.

     

Characterization of lactoferrin binding to the MAC-T bovine mammary epithelial cell line using a biotin-avidin technique

• 1.1. A non-radioisotopic method utilizing a biotin-avidin approach was used to characterize lactoferrin binding to the clonal MAC-T bovine mammary epithelial cell line.
• 2.2. Binding of lactoferrin to MAC-T cells and isolated membranes was specific and saturable.
• 3.3. Unlabeled lactoferrin competed for and displaced biotin-labeled lactoferrin from binding sites on mammary epithelial cells. In contrast, unlabeled transferrin did not compete.
• 4.4. Scatchard analysis of lactoferrin binding to MAC-T cell crude membranes was nonlinear, revealing two classes of binding sites with association constants (K_a) of 2.36 × 10⁷ and 3.36 × 10⁶M⁻¹.
• 5.5. Binding of lactoferrin to MAC-T cells may be associated with the initial events which result in decreased MAC-T cell proliferation.

     

Bioelectrical potentials across the mantle epithelium of Achatina fulica

• 1.1. The shell side of the mantle of Achatina fulica is several millivolts positive to the blood side in vitro.
• 2.2. The electrical potential does not depend on Na⁺, Ca²⁺, Mg²⁺, K⁺ or HCO₃⁻ but requires the presence of chloride on the shell side.
• 3.3. The potential difference and short-circuit current ranged from 3.0 to 30.0 mV and 15.0 to 75 μA/cm² with averages at 10m V and 50 μA/cm² respectively.
• 4.4. The electrical gradient is reduced by 2,4-dinitrophenol, thiocyanate and furosemide but not by ouabain, CO₂ or acetozolamide.
• 5.5. It is suggested that the nature and mechanism of electrogenesis in Achatina parallels that of the Helix mantle.

     

The dynamics of the permeation of an analogue of insect juvenile hormone into nematodes

• 1.1. ¹⁴C-dichlorofarnesoate permeated rapidly into Haemonchus contortus (infective juveniles) and Panagrellus redivivus (mixed cultures) and was strongly bound by hydrophobic association (K_s > 10⁻⁴M).
• 2.2. Uptake rose linearly with increases in temperature (5–38°C) and external concentration (C₀; 0.07–2.15 × 10⁻⁴ M). Within 1 hr the internal concentration, C₁ was >C C₀.
• 3.3. The pH of the medium (6–8) did not affect uptake.
• 4.4. Efflux of dichlorofarnesoate was low: the half-time of release was > 18 hr.
• 5.5. The uptake curve approximated to the expression C₁/C₀ = a(1 − e^−bt) with a and b as constants and t in hr.
• 6.6. These results clarify previous work on the inhibitory action of juvenile hormone on the development of nematodes.

     

The mechanism of interaction of ceruloplasmin with superoxide radicals

• 1.1. The mechanism of interaction of CP with O₂ radicals in chemical and enzymatic systems of Superoxide radical generation as well as in the pulse radiolysis technique was studied.
• 2.2. It is found that CP does not exert any kinetic influence on the decomposition of Superoxide radical and, unlike SOD, cannot catalyze the reaction of disproportionation of these radicals in systems with chemical and enzymatic generation of O₂⁻.
• 3.3. The data obtained confirm the suggestion that CP interacts with precursors of ₂⁻ radicals.
• 4.4. The irradiation of CP does not change its inhibiting activity in the reaction of the formation of Superoxide radicals in systems with enzymatic O₂⁻ generation, but decreases its oxidase activity.
• 5.5. The results obtained demonstrated that the increase in the radiation dose resulted in the decrease of the inhibiting activity of SOD, whereas the activity of CP did not change.

     

Inhibiton and gamma-aminobutyric acid-induced conductance on locust (Schistocerca gregaria) extensor tibiae muscle fibres

• 1.1. Increases in membrane conductance (g_m) were induced by GABA in distal bundles 32, 33 and 34 of extensor tibiae muscles of the locust (Schistocerca gregaria).
• 2.2. Bath application of GABA (10⁻⁵−5 × 10⁻³ M) induced reductions in muscle fibre space constant (λ).
• 3.3. GABA (5 × 10⁻³ M) induced additional membrane conductance of 2.21 ± 0.03 × 10⁻⁶ S/mm, 0.38 ± 0.03 × 10⁻⁶ S/mm and 0.29 ± 0.06 × 10⁻⁶ S/mm on muscle bundles 34, 33 and 32 respectively. The greater sensitivity of muscle fibres in bundle 34 to GABA is due at least in part to a larger number of GABA receptors on bundle 34 muscle fibres.
• 4.4. The decrement of electrotonic potentials in the presence of GABA were measured over distances of both half fibre length and whole fibre length. Good agreement was obtained between changes in space constant produced by GABA using half fibre length and whole fibre length data.
• 5.5. By taking into account changes in space constant induced by GABA it was possible to demonstrate that presynaptic GABA receptors were involved in the inhibition of slow excitatory postsynaptic potentials by GABA.
• 6.6. “Slow” excitatory postsynaptic potentials recorded under current clamp were inhibited in a dose-dependent manner by GABA. This inhibition was not dependent on muscle-fibre GABA sensitivity and could not be completely accounted for by GABA-induced changes in the cable properties of the muscle fibres.

     

Metabolic responses of Cassin's Finches (Carpodacus cassinii) to temperature

• 1.1. The thermal neutral zone of Cassin's Finches extends from 22 to 37.5°C.
• 2.2. Standard metabolism (40.1 Wm⁻² or 7.6kcal bird⁻¹ day⁻¹) of the 28 g birds was 89% of the value predicted for passerines measured at night.
• 3.3. At temperatures below the zone of thermal neutrality metabolism is described by the relation, Wm⁻² = 1.55–74.5°C. The coefficient of heat transfer (1.55Wm⁻²°C⁻¹) is only 58% of the value predicted for birds of this size, indicating excellent insulation.
• 4.4. At temperatures above thermal neutralzfsity metabolism is described by the relation, Wm⁻² = 2.75–62.6°C.
• 5.5. Under conditions of heat stress (44.5°C; P_H2O = 8.6 Torr) Cassin's Finches were able to dissipate up to 208% of their metabolic heat production by evaporative water loss. Maximal rate of water loss was 56 mg g⁻¹ hr⁻¹.
• 6.6. At 20°C resting fasted finches lost a mean of 4.94 ± 1.5 SD mg H₂O g⁻¹hr⁻¹.

     

Charge interactions of cytochrome c with cytochrome c oxidase

• 1.1. The pyridoxal phosphate (PLP) modification of the lysine amino groups in cytochrome c causes decrease in the reaction rate with cytochrome c oxidase.
• 2.2. The rate constants for (PLP);-cyt. c, PLP(Lys 86)-cyt. c, PLP(Lys 79)-cyt. c and native cytochrome c (at pH 7.4, 1=0.02) are 3.6 × 10⁻³'sec-', 5.5 × 10⁻³, 5.2 × 10⁻³-'sec⁻¹ and 9.8 × 10⁻³sec⁻¹, respectively.
• 3.3. In spite of the same positive charge of singly PLP-cytochromes c the reaction between PLP(Lys 86)-cyt. c and cyt. c oxidase exhibits the ionic strength dependence that differs from those of the PLP(Lys 79)-cyt. c.
• 4.4. The rate constants at zero and infinite ionic strength for PLP(Lys 86)-cyt. c is 2-fold less than that for PLP(Lys 79)-cyt. c.
• 5.5. The positively charged cytochrome c lysines 86 and 79 form two from four or five predicted complementary charge interactions with carboxyl groups on cytochrome c oxidase.

     

Properties and distribution of Mg2+-HCO−3-ATPase in brush border membranes isolated from rat small intestine

• 1.1. The small intestine was cut into seven segments and properties and distribution of brush border Mg²⁺-HCO⁻₃-ATPase activity in each segment were examined.
• 2.2. The optimal Mg²⁺ concentration was 1.0 mM.
• 3.3. The optimal HCO⁻₃ concentration was 100 mM in the first (duodenal), 50 mM in the 3rd and 40 mM in the 5th segment, respectively.
• 4.4. The optimal pH value was about 9.0.
• 5.5. l-phenylalanine (above 1 mM) and SCN⁻ (above 50 mM) significantly inhibited both Mg²⁺- and Mg²⁺-HCO⁻₃-ATPase activity.
• 6.6. The enzyme activity was found to be highest in the duodenal segment and then gradually decreased in consecutive segments as well as β-glycerophosphatase, Na⁺-K⁺-ATPase and supernatant carbonic anhydrase.
• 7.7. The functional significance of this ATPase and the relationship with carbonic anhydrase was discussed.

     

Sodium transport in red blood cells of lamprey LAmpetra fluviatilis

• 1.1. Unidirectional Na⁺ influx in lamprey red blood cells was determined using ²²Na as a tracer.
• 2.2. Total Na⁺ uptake and amiloride-inhibitable Na⁺ influx increased in a saturable fashion as a function of external Na⁺ concentration (Na_e).
• 3.3. At 141 mM Na_e, the average value of net Na⁺ influx was 13 ± 1.1 and the amiloride-sensitive Na⁺ influx was 5.3±1.1 mmol/l cells per hr (±SE).
• 4.4. The amiloride-sensitive component of Na⁺ influx was significantly activated by 10⁻⁵ M isoproterenol, by 2 × 10⁻⁵ M DNP, and by cell shrinkage.
• 5.5. Furosemide (1 mM) had no effect on the Na⁺ transport in red cells.
• 6.6. The residual amiloride-insensitive component of Na⁺ transport was a linear function of Na_e in the range of 5–141 mM. This transport seems to be accounted for by simple diffusion.

     

The effect of triiodothyronine on glucose utilization in adipocytes from obese rats

• 1.1. In the presence of insulin, 10⁻⁵ M 3,3',5-triiodothyronine (T₃) treatment for 1/2 hr decreased fatty acid synthesis 35% only in adipocytes from lean rats, whereas at 10⁻¹¹ M through 10⁻⁷M T₃ the obese adipocytes had nearly a 20% increase in fatty acid synthesis.
• 2.2. A 2 hr pretreatment of adipocytes with 10⁻⁹ and 10⁻⁷ M T₃ decreased insulin-stimulated fatty acid synthesis by nearly 20% in both lean and obese adipocytes.
• 3.3. In the absence of insulin, the 2 hr pretreatment with 10⁻⁹ M T₃ resulted in a 45% increase in lean adipocyte fatty acid synthesis, though the obese adipocytes required at least 10⁻⁷ M T₃ for 2 hr to increase the non-insulin-stimulated fatty acid synthesis by 50%.
• 4.4. At 10⁻⁹M T₃ concentrations non-insulin-stimulated fatty acid synthesis was increased by 200% in lean adipose tissue explants, but obese adipose expiants were not significantly affected under these conditions.
• 5.5. The addition of 10⁻⁹ M T₃ plus insulin to the explant media decreased fatty acid synthesis by 35% in both the lean and obese tissues.
• 6.6. The results also imply that the low T₃ status of the obese rat may be contributory to the elevated fatty acid synthesis observed in obese adipocytes.

     

l-lysinamidase from Cryptococcus laurenti 112. Purification and properties

• 1.1. The purified enzyme hydrolyzes the linear l-lysinamide and the cycle amide of l-lysine—l-α-amino-ϵ-caprolactam.
• 2.2. The apparent relative molecular mass is 180,000. The enzyme consists of four subunits and the molecular mass of a single subunit was found to be 47,000.
• 3.3. The coefficient of molecular sedimentation equals 8.3 S, the isoelectric point was determined to be pH 4.3
• 4.4. The enzyme is not a glycoprotein. p-Mercuribenzoate binds 10 SH-groups of the native enzyme molecule and 20 SH-groups in the presence of 0.7% SDS.
• 5.5. pH- optimum for the hydrolysis of l-lysine amides was observed to be 7.5–7.7. The enzyme is strictly dependent on Mn²⁺ and Mg²⁺.
• 6.6. The kinetic parameters for the hydrolysis of l-lysinamide where K_m = 3.8 mM and k_cat = 3000 sec⁻¹ For the hydrolysis of cyclic L-lysinamide K_m = 4.8 mM and k_cat = 2600 sec⁻.

     

Comparative aspects of xanthine dehydrogenase activity of Panstrongylus megistus

• 1.1. Organ extracts from Panstrongylus megistus were assayed for xanthine dehydrogenase (E.C. 1.2.1.37) and xanthine oxidase (E.C. 1.2.3.2) activities.
• 2.2. XDH was present in the fat body, Malpighian tubes, testis and ovary, higher levels being found in the fat body. XO activity was practically absent.
• 3.3. Specific enzyme activity was fairly constant regardless the feeding condition of the insects.
• 4.4. The enzyme was able to oxidize xanthine (K_m = 2.8 × 10⁻⁴ M) and hypoxanthine (K_m = 4.0 × 10⁻⁴ M) but unable to oxidize pterine under the experimental conditions.