The primary structure of three hemoglobin chains from the indigo snake (Drymarchon corais erebennus, Serpentes): First evidence for αD chains and two β chain types in snakes

The hemoglobin of the indigo snake (Drymarchon corais erebennus, Colubrinae) consists of two components, HbA and HbD, in the ratio of 1:1. They differ in both their alpha and beta chains. The amino acid sequences of both alpha chains (alpha(A) and alpha(D)) and one beta chain (betaI) were determined. The presence of an alpha(D)chain in a snake hemoglobin is described for the first time. A comparison of all snake beta chain sequences revealed the existence of two paralogous beta chain types in snakes as well, which are designated as betaI and betaII type. For the discussion of the physiological properties of Drymarchon hemoglobin, the sequences were compared with those of the human alpha and beta chains and those of the closely related water snake Liophis miliaris where functional data are available. Among the heme contacts, the substitution alpha(D)58(E7)His-->Gln is unusual but most likely without any effect. The residues responsible for the main part of the Bohr effect are the same as in mammalian hemoglobins. In each of the three globin chains only two residues at positions involved in the alpha1/beta2 interface contacts, most important for the stability and the properties of the hemoglobin molecule, are substituted with regard to human hemoglobin. On the contrary, nine, eleven, and six alpha1/beta1 contact residues are replaced in the alpha(A), alpha(D), betaI chains, respectively

Dimer-tetramer Transition In Hemoglobins From Liophis Miliaris-i. Effect Of Organic Polyphosphates

1. 1. Hemoglobin from the water-snake Liophis miliaris in the stripped form presents high oxygen affinity of about P50 = 1 mmHg and Hill coefficient of about 1.0 at pH from 6.8 to 8.5. 2. 2. In the presence of ATP such values become P50 = 20 mmHg and nH about 2.0, respectively, at low pH from 6.5 to 7.5. 3. 3. When the pH increases an abrupt decrease of both P50 and nH values occurs falling close to those found for the stripped hemoglobin. 4. 4. Gel-filtration in Sephadex G-100 equilibrated with 0.05 M Tris-HCl buffer containing 1 mM EDTA of the stripped hemoglobin show the presence of only one component of mol. wt of about 32,500 dt similar to the dimer of human hemoglobin A. 5. 5. The deoxy form of the dimer previously treated with ATP and placed on Sephadex column in the same condition but containing 1 mM IHP emerges as tetramer with mol. wt similar to that found for human hemoglobin, i.e. of about 65,000 dt. 6. 6. Results of the multiplicity of the snake hemoglobin, as well as the large alkaline Bohr effect in the presence of ATP previously reported, seems to be inconsistent due to the dimer-tetramer transition that occurs when ATP is bound to the stripped hemoglobin. 7. 7. A molecular mechanism involving the dimer-tetramer transition is proposed to described the oxygen transport in these animals. © 1987.864683687Atha, Riggs, Tetramer-dimer dissociation in hemoglobin and the Bohr effect (1976) J. biol. Chem., 251 (18), pp. 5537-5543Antonini, Brunori, The equilibrium of hemoglobin and myoglobin with ligands (1971) Hemoglobin and Myoglobin in Their Reaction with Ligands, pp. 153-186. , North Holland, AmsterdamChiancone, Gilbert, Dissociation of hemoglobin into subunits. I. Oxyhemoglobin: effect of acetic acid (1965) J. biol. Chem., 240, pp. 3866-3875Douglas, Haldane, Haldane, The laws of combination of haemoglobin with carbon monoxide and oxygen. (1912) J Physiol, 44, p. 275. , ApudSullivan, Riggs, The subunits dissociation properties of turtle hemoglobin (1967) Biochim. biophys. Acta, 140, pp. 274-283Focesi, Jr., Brunori, Bonaventura, Wilson, Galdames-Portus, Effect of pH on the kinetics of oxygen and carbon monoxide reactions with hemoglobins from the airbreathing fish, Loricarüchthys (1979) Comp. Biochem. Physiol., 62 A, pp. 169-171Heatwole, Seymour, Respiration of marine snakes (1976) Respiration of Amphibious Vertebrates, pp. 375-389. , G.M Hughes, Academic Press, LondonLaurent, Killander, A theory of gel filtration and its experimental verification (1964) Journal of Chromatography A, 14, pp. 317-325Matsuura, Ogo, Focesi, Jr., Multiplicity and immunological characterization of the haemoglobin components, from Liophis miliaris and Helicops modestus (1983) Comp. Biochem. Physiol., 76 B, pp. 915-919Monod, Wyman, Changeaux, On the nature of allosteric transitions: a pausible model (1965) J. molec. Biol., 12, pp. 88-118Ogo, Abe, Focesi, Jr., Oxygen dissociation constant in hemoglobin of Helicops modestus and Liophis miliaris, two water-snakes with different morphological adaptations to their environment (1979) Comp. Biochem. Physiol., 63 A, pp. 285-289Ogo, Matsuura, Focesi, Jr., Content of organic polyphosphates and their allosteric effects on haemoglobins from the water-snakes Helicops modestus and Liophis miliaris (1984) Comparative Biochemistry and Physiology Part A: Physiology, 18 A, pp. 587-589Rossi-Fanelli, Antonini, Studies on the oxygen and carbon-monoxide equilibria of human myoglobin (1958) Archives of Biochemistry and Biophysics, 77, pp. 428-492Sekino, Focesi, Bonaventura, Bonaventura, Functional properties of Aphysia brasiliana myoglobin (1978) Comp. Biochem. Physiol., 61 A, pp. 223-226Sullivan, Oxygenation properties of snake hemoglobin (1967) Science, N.Y., 157, pp. 1308-1310Sullivan, Riggs, The subunits dissociation properties of turtle hemoglobin (1967) Biochim. biophys. Acta, 140, pp. 274-28

Functional And Dissociation Properties And Structural Organization Of The Hemocyanin Of Ampullaria Canaliculata (gastropoda, Mollusca)

1. 1. The molecular weight of the native hemocyanin from Ampullaria canaliculata determined by gel filtration has a molecular mass of 7.5 × 106 Da. Dialysis against buffer free of Ca2+ and Mg2+ ions dissociated the hemocyanin into molecular species of 1.0 × 106 and 3.2 × 105 Da. 2. 2. The oxygen-binding curves of hemocyanin were sigmoidal with Hill coefficient of about 2. A positive Bohr effect was demonstrable at pH values ranging from 6.5 to 9.0. 3. 3. Differences between associated and dissociated hemocyanin as well as the effect of the divalent cations on the aggregation of hemocyanin molecule were illustrated by immunoelectrophoresis analysis. 4. 4. Negatively stained preparations of hemocyanin observed by TEM showed the cylindrical configuration of molluscan hemocyanins molecules. Its inner structure, enhanced by the negative contrasting, reveals a regular periodicity as a result of the alignment of serial subunits. © 1993.10503/04/15725730Barros, Propriedades fisico-quimicas e funcionais da hemocianina de Molusco Gastropoda Ampullaria canaliculata (1992) Dissertation Thesis, p. 58Burky, Burky, Buoyancy changes as related to respiratory behavior in an amphibious snail, Pomacea urcelus (Muller), from Venezuela (1977) The Nautilus, 91, pp. 97-104van Bruggen, Schutter, van Breemen, Bijlholt, Wichertjes, Arthropodan and Molluscan hemocyanins (1981) Electron Microscopy of Proteins, 1, p. 1938. , J.R. Harris, Academic Press, Londonvan Bruggen, An electron microscopists view of the quaternary structure of arthropodan and molluscan hemocyanins (1983) Life Chem. (Suppl.), 1, pp. 1-14Cruz-Hofling, Barròs, Matsuura, Focesi, Jr, Preliminary electron microscopic studies of hemocyanin from the Gastropod Ampullaria canaliculata (1992) Proceedings of the 1st Atlantic Congress of Electron Microscopy, , Merida, VenezuelaD'Enghien, Elliot, Bartels, van Bruggen, The haemocyanin of Pila leopoldvillensis—II. Quaternary structure and oxygenation (1971) Comp. Biochem. Physiol., 40 B, pp. 1045-4055Ellerton, Ellerton, Robinson, Hemocyanin—A current perspective (1983) Prog. biophys. molec. Biol., 41, pp. 143-248Fernández-Morán, van Bruggen, Ohtsuki, Macromolecular organization of hemocyanins and apohemocyanins as revealed by electron microscopy (1966) Journal of Molecular Biology, 16, pp. 191-207Hamilton, Herskovits, Furcinitti, Wall, Scanning transmission electron microscopic study of Molluscan hemocyanins in various aggregation states: comparison with light scattering molecular weights (1989) J. ultrastruct. molec. struct. Res., 102, pp. 221-228Heirwegh, Borgion, Lontie, Separation and absorption spectra of alpha and beta-hemocyanin of Helix pomatia (1961) Biochim. biophys. Acta, 48, pp. 517-526Herskovits, Recent aspects of the subunit organization and dissociation of hemocyanins (1988) Comp. Biochem. Physiol., 91 B, pp. 579-611Herskovits, Russell, Carberry, Light-scattering investigations of the subunit structure and sequencial dissociation of Homarus americanus hemocyanin (1984) Biochemistry, 23, pp. 1875-1881Herskovits, Blake, Gonzalez, Hamilton, Wall, Subunit structure and higher order assembly of the hemocyanins of the Melogenidae family: Melongena corona (Gmelin), Bysicon canaliculatum (Linne), B. carica (Gmelin), B. contrarium (Conrad), and B. spiratum (Lamarck) (1989) Comp. Biochem. Physiol., 94 B, pp. 415-421Herskovits, Otero, Hamilton, The hemocyanin of the Ramshorn snail, Marisa cornuarietis (Linne) (1990) Comp. Biochem. Physiol., 97 B, pp. 623-629Lamy, Immunological approach to the structure and evolution of hemocyanins (1982) EMBO Workshop, pp. 15-25. , Life ChemLarimer, Riggs, Properties of hemocyanins—I. The effect of calcium ions on the oxygen equilibrium of crayfish hemocyanin (1964) Comp. Biochem. Physiol., 13, pp. 35-46Laurent, Killander, A theory of gel filtration and its experimental verification (1964) Journal of Chromatography A, 14, pp. 317-330Mangun, Respiratory function of the hemocyanins (1980) Am. Zool., 20, pp. 19-38Mangun, Salt sensitivity of the hemocyanin of euri- and stenohaline squids (1991) Comparative Biochemistry and Physiology Part A: Physiology, 99 A, pp. 159-161Masseyeff, Gombert, Tanguy, Neuzil, Contribution a l'etude biochimique d'hémocyanine de Cymbium neptuni. II. Etude électrophoretique et immunoelectrophoretique (1963) Bull. Soc. chim. Biol., 45, pp. 1133-1144Morse, Meyhofer, Otto, Kuzirian, Hemocyanin respiratory pigment in bivalve molluscs (1986) Science, 231, pp. 1302-1304Petrovich, Morris, McMahon, Oxygen binding by the hemocyanin of Busycon canaliculatum (1990) Comparative Biochemistry and Physiology Part A: Physiology, 97 B, pp. 45-750Terwillinger, Terwilliger, Schabtach, The quaternary structure of molluscan (Helisoma trivolvis) extracellular hemoglobin (1976) Biochim. biophys. Acta, 453, pp. 101-110Truchot, Factors controlling the “in vitro” and “in vivo” oxygen affinity of the hemocyanin in the crab Carcinus maenas (L) (1975) Resp. Physiol., 24, pp. 173-189Wood, Immunochemical properties of the haemocyanins from Buccinum undatum (L) and Neptunea antiqua (L) (1975) Comp. Biochem. Physiol., 52 B, pp. 219-225Wood, Mosby, The hemocyanin from the river snail Viviparus viviparus (L): some properties and subunit structure (1977) Biochem. Soc. Trans., 5, pp. 696-69

Interactions Of Adenosine Triphosphate With Snake Hemoglobins. Studies In Liophis Miliaris, Boa Constrictor And Bothrops Alternatus

The hemoglobins of three snake species: Liophis miliaris, Bothrops alternatus and Boa constrictor present a single ATP binding site per tetramer. The ATP association constant values for the deoxyhemoglobins at pH 7.5 were about KD ≅ 106 M-1 (107 M-1 for B. contrictor), three to four orders of magnitude higher than the respective values for oxyhemoglobin of about KO ≅ 102 M-1. The deoxyhemoglobin constant values markedly decrease as a function of pH, becoming, at pH 8.5, about KD ≅ 103 M-1 whereas for the oxyhemoglobin the constants remain of about the same, KO ≅ 102 M-1, at the pH range studied. The high ATP binding affinity constants, compared to those of human hemoglobin A, were explained from a molecular structural standpoint, considering L. miliaris hemoglobin, whose complete primary sequence is known. Two distinct amino acid residue differences were found in the β-chain, one being Trp (NA3) (more hydrophobic) in the snake hemoglobin which substitutes the Leu (NA3) in human hemoglobin, and the second being Val 101 β (G3) instead of Glu 101 β (G3). The substitutions could provide an un-neutralized, positively charged, residue Lys-104β and, taking into account its high pK value, the pH dependence of ATP binding affinity for the snake hemoglobin would originate from pH-dependent ionization of phosphate groups of the allosteric effector. The physiological implications of the high ATP binding constant, as well as the possible protective role of the nucleotide binding against the effect of high environmental temperatures on the oxygen dissociation curves, are discussed. © 1994.1094701707Ackers, Linked functions in allosteric proteins: an exact theory for the effect of organic phosphates on oxygen affinity of hemoglobin (1979) Biochemistry, 18, pp. 3372-3380Adair, The hemoglobin system—IV. The oxygen dissociation curve of hemoglobin (1925) J. biol. Chem., 63, pp. 529-538Amiconi, Bertollini, Bellelli, Coletta, Condó, Brunori, Evidence for two oxygen-linked binding sites for polyanions indromedary hemoglobin (1985) Eur. J. Biochem., 150, pp. 387-393Antonini, Condó, Giardina, Ioppolo, Bertollini, The effect of pH and d-glycerate-2,3-biphosphate on the O2 equilibrium of normal and SH (β-93) modified human hemoglobin (1982) Eur. J. Biochem., 121, pp. 325-328Bartlett, Phosphate compounds in vertebrate red blood cells (1980) Am. Zool., 20, pp. 103-114Benesch, Edalji, Benesch, The allosteric effect of inositol hexasulfate on oxygen binding by hemoglobin (1976) Biochemistry, 15, pp. 3396-3398Cashon, Bonaventura, Bonaventura, Focesi, Jr, The nicotinamide adenine dinucleotides as allosteric effectors of human hemoglobin (1986) J. biol. Chem., 261, pp. 12700-12705Focesi, Jr, Bonilla, Nagatomo, Matsuura, Dimer-tetramer transition in hemoglobin from Liophis miliaris—III. The phenomenon in snake species of different evolutionary levels (1992) Comp. Biochem. Physiol., 103 B, pp. 985-989Focesi, Jr, Ogo, Matsuura, Dimer-tetramer transition in hemoglobins from Liophis miliaris—II. Evidence with the stripped protein (1990) Comp. Biochem. Physiol., 96 B, pp. 119-122Focesi, Jr, Takagi, Ogo, Polyphosphate binding sites in Liophis miliaris hemoglobin (1990) Evidence with reduced nicotinamide adenine dinucleotide phosphate, 62, pp. 401-408. , Ann. Acad. bras. Ci., (4)Imai, (1992) Allosteric Effects in Haemoglobin, , Cambridge University Press, Cambridge, U.KIslam, Persson, Zaidi, Jornvall, Sea-snake (Microcephalopis gracilis) hemoglobin: primary structure and relationships to other forms (1990) J. Prot. Chem., 9, pp. 533-541McDonald, Turci, Bleichman, Functional and subunit assembly properties of hemoglobin Alberta (α2β2 101Glu→Gly) (1985) J. Mol. Biol., 183, pp. 105-112MacMahon, Hamer, Effect of temperature and photoperiod on oxygenation and other blood parameters of the sidewinder (Crotalus cerastes): adaptative significance (1975) Comp. Biochem. Physiol., 51 A, pp. 59-69Matsuura, Fushitani, Riggs, The amino acid sequences of the α and β chains of hemoglobin from the snake, Liophis miliaris (1989) J. biol. Chem., 264 (10), pp. 5515-5521Matsuura, Ogo, Focesi, Jr, Dimer-tetramer transition in hemoglobins from Liophis miliaris—I. Effect of organic polyphosphate (1987) Comp. Biochem. Physiol., 86 A, pp. 683-687Naqvi, Nadvi, Zaidi, Partial sequence of hemoglobin from cobra (Naja naja naja) (1987) Biosci. Rep., 7, pp. 813-819Ogo, Focesi, Jr, Cashon, Bonaventura, Bonaventura, Interactions of nicotinamide adenine dinucleotides with varied states and forms of hemoglobin (1989) J. Biol. Chem., 264, pp. 11302-11306Oyama, Jr, Nagatomo, Bonilla, Matsuura, Focesi, Jr, Bothrops alternatus hemoglobin components (1993) Oxygen binding properties and globin chain hydrophobic analysis, 105 B, pp. 271-275. , Comp. Biochem. Physiol., (2)Powers, Molecular ecology of Teleost fish hemoglobin: strategies for adapting to changing environment (1980) Am. Zool., 20, pp. 139-162Press, Flannery, Teukolsky, Vetterling, (1986) Numerical Recipes: The Art of Scientific Computing, , Cambridge University Press, Cambridge, U.KRiggs, Wolbach, Sulfhydryl groups and the structure of hemoglobin (1956) The Journal of General Physiology, 39, pp. 585-605Rossi-Fanelli, Antonini, Studies on the oxygen and carbon monoxide equilibria of human hemoglobin (1958) Archs Biochem. Biophys., 77, pp. 478-492Shih, Jones, Imai, Tyuma, Involvement of Glu G3(101)β in the function of hemoglobin (1985) Comparative O2 equilibrium studies of human mutant hemoglobins, 260, pp. 5919-5924. , J. biol. ChemSzabo, Karplus, A mathematical model for structure-function relations in hemoglobin (1972) J. Mol. Biol., 72, pp. 163-197Szabo, Karplus, Analysis of the interaction of organic phosphates with hemoglobin (1976) Biochemistry, 15, pp. 2869-2877Weber, Jensen, Functional adaptation in hemoglobins from ectothermic vertebrates (1988) A. Rev. Physiol., 50, pp. 161-179Wyman, Jr, Linked functions and reciprocal effects in hemoglobin: a second book (1964) Adv. Prot. Chem., 19, pp. 223-28

Bothrops Alternatus Hemoglobin Components. Oxygen Binding Properties And Globin Chain Hydrophobic Analysis

1. 1. Bothrops alternatus oxyhemolysate showed two components by DE-52 cellulose ion-exchange chromatography and polyacrylamide gel electrophoresis: Hb I representing 70% of the hemolysate and Hb II (30%); both are dimeric in the stripped from (mol.wt 32,500 Da) and tetrameric in the presence of IHP (64,000 Da). 2. 2. Hb I, Hb II and whole hemolysate showed functionally similar properties to those of Liophis miliaris, i.e. for the stripped form over the pH interval 7.2-8.9; log P50 values decreasing from ±0.1 to ± -0.15 (thereby an alkaline Bohr effect); ΔH+ = -0.38 and Hill coefficient values decreasing from nH = 1.5 to 1.0. In the presence of ATP, an abrupt decrease in O2 affinity occurs and the log P50 values change from 1.0 to 0.5; the Bohr effect increases to ΔH+ = -0.7 whereas the nH values decrease from ≥2 to values close to unity. 3. 3. For B. alternatus, at a physiological pH range (7.8-9.0) the hemoglobin Bohr effect becomes apparent only in the presence of ATP and this seems to be fundamental for the O2 uptake of the snake. 4. 4. HPLC analysis of the globins shows eight different chains instead of four, as found in L. miliaris hemoglobin, which corroborates the presence of Hb I and Hb II components in B. alternatus, and also shows that the unique tetramer formed from different α and β chains is also consistent in this snake. © 1993.1052271275Antonini, Brunori, (1971) Hemoglobin and Myoglobin in their Reaction with Ligands, , North-Holland, AmsterdamBartlett, Phosphate compounds in vertebrate red blood cells (1980) Am. Zool., 20, pp. 103-114Bonilla, Estrutura, a funçã e evolução em hemoglobinas de ofidios (1992) Ph.D. Thesis, , Universidade Estadual de Campinas, BrazilDessauer, Fox, Ramirez, Preliminary attempt to correlate paper-electrophoretic migration of hemoglobin with phylogeny in Amphibia and Reptilia (1957) Archs Biochem. Biophys., 71, pp. 11-17Focesi, Jr, Bonilla, Nagatomo, Matsuura, Dimer-tetramer transition in hemoglobin from Liophis miliaris. III. The phenomenon in snake species of different evolutionary levels (1992) Comp. Biochem. Physiol., 103 B, pp. 985-989Focesi, Jr, Ogo, Matsuura, Dimer-tetramer transition in hemoglobins from Liophis miliaris. II. Evidence with the stripped protein (1990) Comp. Biochem. Physiol., 96 B, pp. 119-122Matsuura, Ogo, Focesi, Jr., Dimer-tetramer transition in hemoglobins from Liophis miliaris. I. Effect of organic polyphosphate (1987) Comp. Biochem. Physiol., 86 A, pp. 683-687Matsuura, Fushitani, Riggs, The amino acid sequences of the α and β chains of hemoglobin from the snake, Liophis miliaris (1989) J. biol. Chem., 264, pp. 5515-5521Matsuura, Rosa, Greene, Focesi, Jr, Amino acid sequence of the β-chain of heuroglobin from the snake Liophin miliaris (1992) FASEB J., 6 (1), p. AS7Monod, Wyman, Changeaux, On the nature of allosteric transitions: a plausible model (1965) J. molec. Biol., 12, pp. 88-118Ornstein, Disc electrophoresis background and theory (1964) Ann. N.Y. Acad. Sci., 121, pp. 321-349Rossi-Fanelli, Antonini, Studies on the oxygen and carbon-monoxide equilibria of human hemoglobin (1958) Archs Biochem. Biophys., 77, pp. 478-492Schwantes, Hemoglobinas e haptoglobinas em serpentes (1972) Ph.D Thesis, , Universidade Federal do Rio Grande do Sul, BrasilSode, Oxygen binding characteristic of whole-blood and hemoglobin from the snake Thamnophis sirtalis (1991) Comp. Biochem. Physiol., 100 B, pp. 697-703Sullivan, Reptilian hemoglobins (1974) Chemical Zoology, 9, pp. 377-398. , M. Florkin, B.T. Scheer, Academic Press, New York, Amphibia and Reptili