Analysis of oxygen binding by Xenopus laevis hemoglobin: implications for the Root effect.
We have measured oxygen binding properties for red cell suspensions and stripped hemolysate of Xenopus laevis (XL) in order to answer the question whether XL hemoglobin exhibits a Root effect. The present results show that under physiological conditions XL red cells do not exhibit all the criteria for a Root effect. Compared to human Hb A, stripped XL hemoglobin has a low oxygen affinity, a normal alkaline Bohr effect and a lower interaction with 2,3-diphosphoglycerate, its physiological allosteric effector in red cells. The values for the Hill coefficients are lower than those for human hemoglobin but XL Hb remains cooperative (n50 approximately 2), even at pH values below 6. Attempts to mimick some of the criteria for the Root effect were carried out in pure Hb A solution upon addition of potent allosteric effectors. This leads to low cooperativity and less than 100% oxygen saturation under room air at acidic pH. Under these conditions, mammalian or XL Hb have a maximum proton release at pH approximately 8 and an increased reverse Bohr effect. By contrast, a Root effect Hb exhibits a maximum proton release at neutral pH with the absence of reverse Bohr effect. Therefore the Root effect in fish Hb and the extreme stabilization of the T-state with effectors in mammalian Hb are not an identical phenomenon. Without crystallographic analyses of fish Hb exhibiting a Root effect, the molecular interpretation of this functional property still remains unexplained.
Kister
J
J
INSERM Unité 299, Hôpital de Bicêtre, Le Kremlin-Bicêtre, France.
Bohn
B
B
Marden
M C
MC
Poyart
C
C
D000818:Animals; D004912:Erythrocytes / Q000378:metabolism; D006441:Hemoglobin A / Q000378:metabolism; D006454:Hemoglobins / Q000378:metabolism*; D006461:Hemolysis; D006863:Hydrogen-Ion Concentration; D010100:Oxygen / Q000097:blood*; D010108:Oxyhemoglobins / Q000378:metabolism; D014982:Xenopus laevis / Q000097:blood*
D006454:Hemoglobins; D010108:Oxyhemoglobins; D006441:Hemoglobin A; D010100:Oxygen