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The oxygen binding properties of hemoglobin and some hematological parameters in Eskimo dogs (belonging to Canis lupus familiaris) in Ilulissat/Jacobshavn, Greenland were analysed. The average [2,3-DPG] and [Hb] (n = 16) were 3.14 +/- 0.34 mmol l-1 blood and 9.53 +/- 0.65 g dl-1 (1.49 mmol l-1), respectively, giving a stoichiometric ratio of 2.11 mol 2,3-DPG/mol Hb. Oxygen binding analysis carried out on hemolysate in HEPES buffer at 20 and 37 degrees C revealed a high oxygen affinity (1.2 mmHg at pH 7.4, 20 degrees C) in the desalted condition, which decreased markedly in the presence of chloride and 2,3-DPG. A low apparent equilibrium constant for the binding of 2,3-DPG (1.0 x 10(-5) mol l-1) was found at pH 7.2 and 20 degrees C in the absence of chloride. Moreover, we show that chloride ions have an additive effect on oxygen affinity in the concentration range 10-300 mmol l-1 in the presence of 3 mmol l-1 2,3-DPG at low pH and temperature (pH < 7.4 and 20 degrees C). This feature may be of physiological importance to oxygen unloading under acidotic conditions when tissue temperature is low. Thermodynamic analysis reveal that in the presence of 3 mmol l-1 2,3-DPG and 100 mmol l-1 chloride, the Eskimo dog hemoglobin exhibits a low heat of oxygenation, which places this animal close to arctic ruminants with respect to the influence of temperature on oxygen binding in vivo.

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Hemoglobin (Hb) from the Eskimo dog (belonging to Canis lupus familiaris) showed similar Bohr effect (delta log P50/delta pH) to human HbA in the presence of 100 mmol l-1 NaCl at 20 degrees C. The presence of 7% carbon dioxide in the desalted condition caused a positive (reversed) Bohr effect in the pH range 7.1-7.5 on Eskimo dog Hb, whereas in human HbA there was no Bohr effect within this pH range. A positive Bohr effect on Eskimo dog Hb in this condition was also observed at 37 degrees C. This could indicate differences in the pK values of the amino terminal residues of the two hemoglobins, with possible pH-dependent binding of both bicarbonate (HCO(3)-) and carbamate. Analysis of the effect of CO2 on oxygen affinity of Eskimo dog Hb in the pH range 6.7-7.6 in the presence of chloride and/or 2,3-diphosphoglycerate (2,3-DPG) support this theory. Our results indicate a competition between HCO(3)- and Cl- in affecting oxygen binding. Thermodynamic analysis reveals that bicarbonate binding lowers the apparent heat of oxygenation in Eskimo dog Hb nearly as much as chloride does in the presence of 2,3-DPG at physiological pH. This safeguards an effective oxygen unloading at lowered red blood cell concentrations of chloride. Moreover, we show that the oxygen affinity at high O2 saturation is less dependent on temperature in the presence than in the absence of CO2-.

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The hemoglobin from the European frog, Rana temporaria, consists of one major and three minor components. The tetramers aggregate upon deoxygenation notably at pH 7:3. Aggregation due to formation of disulphide bridges, as occurs in related species, was observed only in polyacrylamide gels. The hemolysate showed a pronounced Bohr effect. Oxygen affinity decreased with increasing hemoglobin concentration, indicating that aggregation affects the functional properties of the hemolysate. Oxygen binding equilibria of unfractionated hemolysate are insensitive to chloride and show low sensitivity to ATP. Analysis of oxygen equilibrium curves in terms of the two-state allosteric model (MWC) showed that pH change exerted a greater effect on the association constant of the oxygenated state (KR) than that of the deoxy state (KT). The number of interacting binding sites (q) increased with hemoglobin concentration. Cooperativity of oxygen binding, evaluated as Hill coefficient n, never exceeded the value of 3.0. Earlier studies on hemoglobin and blood from this and related species, report significantly higher n values at high O2 saturation. Molecular adaptive aspects are discussed.

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The functional properties of haemoglobin from the Lesser Rorqual whale (Balaenoptera acutorostrata) have been characterized as a function of the heterotropic effector concentrations and temperature. The results obtained suggest the existence of sophisticated modulation mechanisms based on the interplay of organic phosphates, carbon dioxide, lactate and temperature. These, together with the very small apparent heat of oxygenation (delta H) of oxygen binding, have been physiologically interpreted on the basis of the specific metabolic needs of this diving mammal.

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The most surprising characteristic of reindeer hemoglobin (Hb) concerns its response to changes in temperature. Thus, the shape of the oxygen-binding curve is strongly temperature dependent due to the difference in the enthalpy of oxygenation between the T and R state of the molecule. In fact, delta H of oxygen binding to the T state is strongly exothermic whereas that of the R state is very close to zero or possibly positive after correction for the heat of oxygen solubilization. Moreover, the allosteric transition T0----R0 has been found to display a negative delta H and a contemporaneous decrease in entropy, a behavior which is precisely the opposite of what has been reported for other hemoglobins. As a whole, reindeer Hb represents a beautiful example of the significance that comparative studies may have in assessing the general validity of the main properties of the hemoglobin molecule.

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The oxygen binding of whole blood from humans and two arctic mammals, reindeer and muskox, has been studied as a function of carbon dioxide and temperature. All bloods display a marked Bohr effect with Bohr coefficients in the range -0.44- -0.73. The Bohr effect is more pronounced at 20 degrees C. The temperature sensitivity of reindeer and muskox blood expressed by the apparent heat of oxygenation, delta H, is almost three times lower than that of human HbA under the same experimental conditions. This thermodynamic difference gives special benefits to arctic mammals with large heterothermy by safeguarding oxygen unloading at very low ambient temperatures.

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Previous results [(1988) Arct. Med. Res. 47, 83-88] have shown that hemoglobin from reindeer is characterized by a low overall heat of oxygenation. This particular aspect has been investigated further in a series of precise oxygen equilibrium experiments. The results obtained show a peculiar dependence of the temperature effect on the fractional saturation of hemoglobin with oxygen, which could be regarded as a very interesting case of molecular adaptation to extreme environmental conditions.

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1. The functional properties of hemoglobin from the reindeer (Rangifer tarandus tarandus L.) are characterized as a function of pH, temperature and organic phosphate concentration. 2. Alongside overall similarities shared with most vertebrate hemoglobins, hemoglobin from the reindeer shows significant differences with respect to the effect of both organic phosphates and chloride anions. 3. The limited effect of temperature on oxygen binding (delta H = -4 kcal/mol O2) could be regarded as an interesting case of molecular adaptation to extreme environmental conditions.

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1. The hemoglobin system from musk ox (Ovibos muschatos) has been characterized from the functional point of view with special regard to the effect of organic phosphates and temperature. 2. The results are similar to those previously obtained in the case of reindeer and confirm that hemoglobins from arctic animals may display very low enthalpy change for the reaction with oxygen. 3. This finding is considered an example of molecular adaptation of respiratory pigments to extreme environmental conditions.

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