Martha Farmer

Occurrence of root effect hemoglobinsin Amazonian fishes

Abstract o 1. The Root effect was measured in hemolysates from representatives of 56 genera of Amazonianfishes. 2. Hemolysates from several species of air-breathing fishes were found to have Root effects, contraryto published hypotheses. 3. Hemolysates from Potamotrygon, a freshwater ray, exhibit a Root effect under our experimental conditions. 4. The pattern of Root effect distribution correlates positively with the distribution of choroid retiamirabile and swimbladders, but not with the distribution of swimbladder retia mirabile; it is proposed that the former is the more primitive structure which is associated with the origin of Root effect hemoglobins 5. Some of the fish hemoglobins differ spectrally from one another. The positions of the absorptionmaxima of the deoxyhemoglobins range from 553 nm (Lepidosiren paradoxa and Potamotrygon sp.) to 560 nm (Plagioscion) 6. Occurrence of Root effects is not correlated with the complexity of the hemoglobin electrophoreticpattern, although several species are found to have multiple hemoglobin systems in which the Root effect is restricted to certain components.

The transition from water to air breathing:Effects of CO2 on hemoglobin function*

Abstract o 1. The pH dependence of the specific effect of CO2 (pCo2 = 30 torr) on oxygen equilibria has been measured for the hemoglobins of four species of fish and one amphibian. Leiostomus xanthurus (marine teleost). Brachyplatystoma sp. (Amazonian catfish), Hoplosternum littorale (air-breathing catfish), Lepidosiren paradoxa (air-breathing lungfish), and Typhlonectes compressicauda (a caecilian). 2. The blood CO2 content of air-breathing fish and amphibians is considerably higher than that of water breathers, yet the hemoglobins examined from the air breathers showed no special adaptation to the increased CO2 load. 3. Although the effect of pH on the oxygen equilibria of the different hemoglobins varied greatly, from Root effect to reverse Bohr effect, the effect of CO2 on the oxygen affinity was very similar for all but that of the catfish, Brachyplatystoma sp., for which the effect was somewhat larger. 4. The drop in oxygen affinity brought about by CO2 increased with increasing pH for each hemoglobin examined, and was about half that produced by CO2 for human Hb A. This suggests that half the chains of each hemoglobin may have blocked amino terminal groups, except possibly Brachyplatystona s.p. hemoglobin.

A fetal-maternal shift in the oxygen equilibrium of hemoglobin from the viviparous caecilian,Typhlonectes compressicauda

1. The equilibria and kinetics of oxygen binding by blood and hemoglobin from adult and fetal caecilians,Typhlonectes compressicauda, have been measured. 2. The oxygen affinity of fetal blood is higher than that of adult blood. 3. Electrophoresis of adult and fetal hemoglobins suggests that they may be identical: a major and minor component occurs in each. 4. Adult and fetal hemoglobins have identical oxygen equilibria. Stripped hemoglobins have a high oxygen affinity and no Bohr effect between pH 6.5 and 10.0. An “acid”, reversed Bohr effect is present below pH 6.5. The addition of 1 mM ATP reduces the oxygen affinity markedly and produces a moderate, normal Bohr effect. 5. The major nucleoside triphosphate in fetal and adult erythrocytes is adenosine triphosphate: about 10% of the nucleoside triphosphates is guanosine triphosphate. Adult erythrocytes contain 3 times as much ATP as do the fetal erythrocytes. 6. The fetal to maternal shift in the oxygen equilibrium is mediated entirely by the difference in ATP content of the maternal and fetal red blood cells.

Equilibria and kinetic of oxygen and carbon monoxide binding to the haemoglobin of the South American lungfish,Lepidosiren paradoxa

Abstract 1. The haemoglobin of the South American lungfishLepidosiren paradoxa has a single component. 2. The equilibria of this respiratory protein with oxygen have been investigated both in the blood and with the purified haemoglobin. There is a substantial, normal, alkaline Bohr effect and marked sensitivity to organic phosphates in the haemoglobin solutions. 3. Studies on the pH dependence of the kinetics of oxygen dissociation can be interpreted in terms of a normal Bohr effect. 4. The kinetics of combination of carbon monoxide have an unusual pH dependence. 5. These findings are discussed in terms of the two-state model of Monodet al. (1965)

Functional properties of hemoglobin and whole blood in an aquatic mammal, the Amazonian manatee (Trichechus inunguis)

Abstract 1. Hematocrit (43%) and O 2 binding capacity (18.8 ml O 2 /100 ml blood) of Trichechus inunguis blood are low compared to the values for other diving mammals but are similar to those for land mammals. 2. Stripped manatee Hb is similar to human Hb A in its sensitivity to pH, 2,3-diphosphoglycerate and CO 2 , but less sensitive to temperature and more prone to dissociate into dimers. 3. The unique Hill plots exhibit no cooperativity below 30% O 2 -saturation indicating a highly stabilized T or low-affinity state(s); such asymmetric Hill plots together with biphasic O 2 -binding kinetics could mean chain heterogeneity. 4. The pH dependence of oxygen binding by the apparent T state, hemoglobin as seen in the Hill plots, is enhanced by 2,3-diphosphoglycerate but eliminated by CO 2 .

Studies of the functional properties of the hemoglobins ofOsteoglossum bicirrhosum andArapaima gigas

Abstract 1. The effects of pH and organic phosphate on the equilibrium and kinetic properties of the binding of ligands to the hemoglobins of two related Amazonian fish, Osteoglossum bicirrhosum , an obligate water breather, and Arapaima gigas , an obligate air breather, have been studied. 2. The hemoglobins of both fish exhibit a Root effect, and the minimum oxygen affinity is associated with the complete loss of cooperative ligand binding. 3. In this low affinity, T state, there is great subunit heterogeneity in both hemoglobins as evidenced by Hill coefficients well below unity and biphasic carbon monoxide combination kinetics. 4. The greatest difference in the properties of the hemoglobins of these two fish is found at high pH (above pH 8) where both hemoglobins exhibit their highest ligand affinities. Here the hemoglobins differ both in the affinity and cooperatively with which they bind ligands, Arapaima hemoglobin having the lowest affinity but the highest cooperativity.

Ocorrência de hemoglobinas de efeito Root em peixes amazônicos

O efeito Root foi medido em hemolisados de representantes de 56 gêneros de peixes amazônicos. Hemolisados de varies espécies de peixes de respiração aérea apresentaram efeito Root, contrariamente às hipóteses publicadas. Hemolisados de Potamotrygon, uma arraia de água doce, exibiram efeito Root sob nossas condições experimentais. O padrão de distribuição do efeito Root correlaciona-se positivamente com a distribuição das retia mirabile da coróide e a distribuição das bexigas natatorias, mas não com a das retia mirabile da bexiga natatoria; propõe-se que a primeira é a estrutura mais primitiva que está associada à origem das hemoglobinas com efeito Root. Algumas das hemoglobinas de peixe diferem espectralmente uma das outras. As posições do máximo de absorção das desoxihemogJobinas vão de 553 nm (Lepidosiren paradoxa e Potamotrygon sp.) a 560 nm (Plagioscion). Ocorrência de efeitos Root não está correlacionada com a complexidade do padrão eletroforético da hemoglobina, embora em várias espécies se tenha encontrado sistemas de hemoglobina múltipla, nos quais o efeito Root está restrito a certos componentes.