High affinity and temperature sensitivity of blood oxygen binding in Pangasianodon hypophthalmus due to lack of chloride-hemoglobin allosteric interaction

Air-breathing fishes represent interesting organisms in terms of understanding the physiological changes associated with the terrestrialization of vertebrates, and, further, are of great socio-economic importance for aquaculture in Southeast Asia. To understand how environmental factors, such as high temperature, affect O₂ transport in air-breathing fishes, this study assessed the effects of temperature on O₂ binding of blood and Hb in the economically important air-breathing fish Pangasianodon hypophthalmus. To determine blood O₂ binding properties, blood was drawn from resting cannulated fishes and O₂ binding curves made at 25°C and 35°C. To determine the allosteric regulation and thermodynamics of Hb O₂ binding, Hb was purified, and O₂ equilibria were recorded at five temperatures in the absence and presence of ATP and Cl^-. Whole blood had a high O₂ affinity (O₂ tension at half saturation P₅₀ = 4.6 mmHg at extracellular pH 7.6 and 25°C), a high temperature sensitivity of O₂ binding (apparent heat of oxygenation ΔH_app = -28.3 kcal/mol), and lacked a Root effect. Further, the data on Hb revealed weak ATP binding and a complete lack of Cl^- binding to Hb, which, in part, explains the high O₂ affinity and high temperature sensitivity of blood O₂ binding. This study demonstrates how a potent mechanism for increasing O₂ affinity is linked to increased temperature sensitivity of O₂ transport and provides a basic framework for a better understanding of how hypoxia-adapted species will react to increasing temperatures.