Effects of buffer composition, pH and temperature on oxygen binding by planorbid snail haemoglobins

Abstract

The oxygen-binding properties of the haemoglobins of four species of freshwater planorbid snails Bulinus africanus, Biomphalaria glabrata, Bulinus tropicus and Heliosoma trivolvis were examined. Samples chromatographed on Sephadex G75 in CO-saturated buffer (0,1 mol dm⁻³ MgCl₂, 0,05 mol dm⁻³ Tris-HCl, 0,5 mmol dm⁻³ phenylmethyl sulfonyl fluoride (PMSF), 1,0 mmol dm⁻³Dithioerytritol, ionic strength 0,3) gave the best stability against methaemoglobin formation. The highest haemoglobin oxygen affinity (P₅₀ = 0,35 mmHg) was measured with MgCl₂-buffer with a Hill coefficient, n_max, of 1,16. An n_max value of 2,2 was obtained with snail buffer (4 mmol dm⁻³ KCl, 50 mmol dm<⁻³ NaCl, 1 mmol dm⁻³ MgCl₂, 6 mmol dm⁻³ CaCl₂, 50 mmol dm⁻³ Tris-HCI, 0,5 mmol dm⁻³ PMSF, ionic strength 0,075, pH 8,0) that gives a P₅₀ value of 1,58 mmHg. A Bohr effect of between −0,44 and −0,11 was measured in the physiological range of pH 8,0–7,1 in snail buffer for the three species, B. glabrata, B. africanus and B. tropicus. For these planorbids the heat of oxygenation of the haemoglobins, ΔH, was −95,3 kJ/mol and −68,1 kJ/mol at pH 8,0 and pH 7,1 respectively.