The hemoglobin of the common sting-ray, Dasyatis sabina: Structural and functional properties

doi:10.1016/0305-0491(78)90129-3

Comparative Biochemistry and Physiology Part B: Comparative Biochemistry

Volume 60, Issue 2, 1978, Pages 189-193

https://doi.org/10.1016/0305-0491(78)90129-3 Get rights and content

Abstract

1.
1. The hemoglobin of the sting-ray, Dasyatis sabina, is both polymorphic and heterogeneous; three components predominate.
2.
2. One major component has two kinds of polypeptide chain, of which one, presumably an α-chain, has a blocked NH₂-terminus and an arginyl COOH-terminus, whereas carboxypeptidases A and B release tyrosine and histidine from the COOH-terminus of the β-chain.
3.
3. The amino acid sequence of the beginning NH₂-terminal segment of the β-chain of the major component has been determined.
4.
4. The hemoglobin of the sting-ray, Dasyatis sabina, is highly resistant to urea and does not dissociate readily into subunits.
5.
5. Oxygen binding by the hemoglobin is not affected by organic phosphates or high concentrations of either NaCl or urea.
6.
6. The hemoglobin does not polymerize beyond tetramers.
7.
7. Cooperativity, as monitored by n in the Hill equation, is pH-dependent and maximal between pH 8.5 and 9.0.
8.
8. The hemoglobin has a large Bohr effect; the oxygen affinity is 16 times higher at pH 10 than at pH 6.5.

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D.H. Atha et al.
Tetramer-dimer dissociation and the Bohr effect
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T.O. Baldwin et al.
The hemoglobins of the bullfrog, Rana catesbeiana, partial amino acid sequence of the β-chain of the major adult component
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Hemoglobin of the electric Atlantic torpedo, Torpedo nobiliana: A cooperative hemoglobin without Bohr effects
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Subunit dissociation in fish hemoglobins
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Structure and function of the hemoglobins of the carp, Cyprinus carpio
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(1972)
R.G. Gillen et al.
The hemoglobins of a freshwater teleost, Cichlasoma cyanoguttatum (Baird and Girard). II. Subunit structure and oxygen equilibria of the isolated components
Archs. Biochem. Biophys.
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C. Manwell
Fetal and adult hemoglobins of the spiny dogfish Squalus suckleyi
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(1963)
R.R. Pennelly et al.
Equilibria and ligand binding kinetics of hemoglobins from the sharks, Prionace glauca and Carcharhinus milberti
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(1975)
C. Tanford
Protein denaturation. Part C. Theoretical models for the mechanism of denaturation
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J. Bonaventura et al.

Urea tolerance as a molecular adaptation of elasmo-branch hemoglobins

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Conformational parameters for amino acids in helical, β-sheet, and random coil regions calculated from proteins

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Cited by (23)

Oxygen and Carbon Dioxide Transport in Elasmobranchs
2015, Fish Physiology
- 1.
  Introduction
- 2.
  Blood-Oxygen Transport
  2.1.
  Hemoglobin
  2.2.
  Red Blood Cell Function and Homeostasis
  2.3.
  Blood-Oxygen Content: Hemoglobin Concentration and Hematocrit
  2.4.
  Whole-Blood-Oxygen Equilibria
- 3.
  Transport and Elimination of Carbon Dioxide
  3.1.
  Carbon Dioxide Transport in Blood
  3.2.
  Carbon Dioxide Excretion
- 4.
  Conclusions and Perspectives
The elasmobranchs are an ecologically diverse subclass of over 1000 species that have evolved to inhabit a wide range of environments and become one of the most speciose groups of vertebrate predators on Earth. This chapter reviews what is known about elasmobranch O₂ uptake, transport, and delivery, as well as CO₂ transport and elimination, and focuses upon two metalloproteins central to these processes, hemoglobin (Hb) and carbonic anhydrase (CA), both of which have undergone distinct functional adaptations. Furthermore, adaptations in relation to life history, which include exercise, hypoxia, salinity, temperature, and, in some species, regional heterothermy, are reviewed. While processes and principles of gas transport and exchange in elasmobranchs are often similar to those of the better described teleosts, there are differences that stand out as clearly worthy of further investigation. Generally, elasmobranch Hbs exhibit a high affinity for O₂ relative to teleosts, which may be associated with a low organic phosphate/Hb ratio and an antagonistic effect of urea on Hb-ATP sensitivity. The Hbs also exhibit a moderate Bohr and Haldane effect, but high buffering by Hb and plasma proteins coupled with the presence of plasma accessible CA greatly reduces the interaction between O₂ and CO₂ exchange relative to the situation in teleosts. Moreover, at least in the dogfish, Squalus suckleyi, current models of CO₂ excretion suggest similar contributions of the plasma and red blood cell (RBC) to CO₂ excretion, a model that contrasts with the pattern of CO₂ excretion typical of other vertebrates in which near exclusive reliance is placed on the RBC. High plasma buffering and plasma-accessible CA in the gill of dogfish favor HCO₃⁻ dehydration in the plasma, while HCO₃⁻ dehydration via the RBC is constrained by low RBC CA activity and the absence of a Haldane effect in this species. In the Hb of the whip stingray, Dasyatis akajei, a novel Bohr effect mechanism has been discovered and this same species possesses a novel ATP binding site in Hb. Finally, in the high performing regionally heterothermic sharks, there appears to be a reduction or reversal in Hb temperature sensitivity consistent with regionally heterothermic teleosts, but this remains to be investigated in detail. While gas transport and exchange is a central process associated with the success of elasmobranchs, it has been most thoroughly investigated in just a few species; clearly a great deal remains to be discovered to achieve a more representative understanding of gas transport and exchange in elasmobranch fish.
Organic phosphates in the red blood cells of fish
2000, Comparative Biochemistry and Physiology - A Molecular and Integrative Physiology
Fish are dependent on aerobic metabolism. They respond to changes in oxygen availability by a wide spectrum of compensatory and respiratory adjustments to safeguard tissue oxygenation. Such adjustments are directed to facilitate both oxygen uptake at the gas exchange surfaces and oxygen unloading to tissues. The importance of erythrocytic organic phosphates as regards oxygen transfer has been recognised since 1967 when the ‘dramatic’ effect of 2,3DPG on human haemoglobin was first reported. The present review examines the appearance of all the major erythrocytic organic phosphates during the evolutionary radiation of fish. In addition, it provides examples illustrating qualitative and quantitative ontogenetic changes of organic phosphates in the red blood cell of several fish species and describes their effects on oxygen affinities. The interaction of the organic phosphates with haemoglobins and divalent cations are also examined. Of particular interest is the regulation of erythrocytic organic phosphates according to both environmental and physiological conditions.
Asymmetric hemoglobins, their thiol content, and blood glutathione of the scalloped hammerhead shark, Sphyrna lewini
1997, Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
Starch gel electrophoresis pH 8.6, or PAGE pH 8.9, of the scalloped hammerhead shark hemolysates showed three hemoglobins (Hb). An additional Hb between the two most mobile electrophoretic components was seen in starch gel electrophoresis, pH 8.1, and also in highly loaded PAGE gels. The relative concentration of these Hbs was variable among individuals, when accessed at pH 8.1. Dilution of hemolysates led to a redistribution of the Hb tetramer subunits. Under denaturing conditions, the unfractionated hemolysate was resolved in 3 Hb subunits. Isolated Hbs, named SL I–SL IV, showed unusual subunit compositions: SL I, the least mobile, is “b₃c”; SL II is “a₂bc”; SL III and SL IV are composed only by “a” subunits. Hemoglobins in the whole hemolysate have an average of two reactive cysteines per tetramer, which were not easily S-thiolated by glutathione, as is the case for related species. After hemoglobin denaturation, six additional -SH groups were titrated by Ellman's reagent. Methemoglobin content was low in the erythrocytes of nine examined specimens, 1.13 ± 1.90%. High values for total erythrocyte glutathione (GSH) were found: 4.5 ± 0.7 mM; n = 7. The ratio of 1.4 ± 0.4 GSH/Hb is higher than usually reported for mammalians.
Multiple hemoglobins in the electric ray: Torpedo marmorata
1996, Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
The elasmobranch Torpedo marmorata has multiple hemoglobins. Some specimens have at least 6–8 components, others 10 or 12–13. We did not observe hemoglobins of molecular weight greater than tetramers. The analyses of hemoglobins polypeptide composition seem to indicate that each hemoglobin contains more than two types of globin chains. We show evidence for the existence of hybrid hemoglobins. Post-translational modifications of globin chains could occur. All these phenomena can contribute to the multiplicity of hemoglobins
High hemoglobin mixed disulfide content in hemolysates from stressed shark
1990, Comparative Biochemistry and Physiology -- Part B: Biochemistry and
- 1.
  1. Hemolystate from heavily stressed smooth hammerhead shark, Sphyrna zygaena, shows three electrophoretic components, SZ I, SZ II and SZ III, whose relative concentrations are 36.4 ± 6.8, 36.4 ± 5.0 and 20.8 ± 5.7%, respectively. After reduction with DTE only SZ I remained.
- 2.
  2. SZ I reacted with glutathione disulfide reconstitute SZ II and SZ III.
- 3.
  3. Non-reduced, DTE-reduced, and denatured hemoglobin were found to have 2.0 ± 0.4, 3.7 ± 0.6, and 9.4 ± 0.7-SH groups, respectively.
- 4.
  4. Erythrocyte non-protein—SH (NPSH), including glutathione present as mixed disulfide with SZ II and SZ III, is 1.7 NPSH/Hb.
Oxygen binding properties of blood and hemoglobin solutions in the carpet shark (Cephaloscyllium isabella): Roles of ATP and urea
1984, Comparative Biochemistry and Physiology -- Part A: Physiology
- 1.
  1. The whole-blood oxygen equilibrium curve in carpet sharks (Cephaloscyllium isabella) at 15°C is weakly cooperative (Hill's coefficient n = 1.5) with a moderate Bohr effect (Δlog $P_{50} Δ pH = −0.32$ ) and high O₂ affinity (P₅₀ = 8.3 mmHg at pH 7.7).
- 2.
  2. Warm acclimated sharks (15°C) did not alter the erythrocytic trinucleotide concentration but had high urea levels which increased O₂ affinity by comparison with sharks acclimated at 5°C.
- 3.
  3. ATP decreased the O₂ affinity of “stripped” Hb solutions whereas urea increased it. Equimolar concentrations of both cofactors suggested suppression of the phosphate effect by urea.
- 4.
  4. Blood from embryonic carpet sharks (12–15 mm long) showed similar O₂ affinities to blood from adult sharks.

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^∗: This work was carried out during the tenure of an NIH post-doctoral fellowship.

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Comparative Biochemistry and Physiology Part B: Comparative Biochemistry

Abstract

References (24)

Carboxypeptidases A and B

Meth. Enzym.

Tetramer-dimer dissociation and the Bohr effect

J. biol. Chem.

The hemoglobins of the bullfrog, Rana catesbeiana, partial amino acid sequence of the β-chain of the major adult component

J. biol. Chem.

Hemoglobin of the electric Atlantic torpedo, Torpedo nobiliana: A cooperative hemoglobin without Bohr effects

Biochim. biophys. Acta

Subunit dissociation in fish hemoglobins

J. biol. Chem.

Structure and function of the hemoglobins of the carp, Cyprinus carpio

J. biol. Chem.

The hemoglobins of a freshwater teleost, Cichlasoma cyanoguttatum (Baird and Girard). II. Subunit structure and oxygen equilibria of the isolated components

Archs. Biochem. Biophys.

Fetal and adult hemoglobins of the spiny dogfish Squalus suckleyi

Archs. Biochem. Biophys.

Equilibria and ligand binding kinetics of hemoglobins from the sharks, Prionace glauca and Carcharhinus milberti

Comp. Biochem. Physiol.

Protein denaturation. Part C. Theoretical models for the mechanism of denaturation

Adv. Protein Chem.

Urea tolerance as a molecular adaptation of elasmo-branch hemoglobins

Science

Conformational parameters for amino acids in helical, β-sheet, and random coil regions calculated from proteins

Biochemistry

Cited by (23)

Oxygen and Carbon Dioxide Transport in Elasmobranchs

Organic phosphates in the red blood cells of fish

Asymmetric hemoglobins, their thiol content, and blood glutathione of the scalloped hammerhead shark, Sphyrna lewini

Multiple hemoglobins in the electric ray: Torpedo marmorata

High hemoglobin mixed disulfide content in hemolysates from stressed shark

Oxygen binding properties of blood and hemoglobin solutions in the carpet shark (Cephaloscyllium isabella): Roles of ATP and urea