| Abstract: | Formation of the binary complex between the reduced coenzyme nicotinamide adenine dinucleotide (NADH) and pig skeletal muscle lactate dehydrogenase (LDH, EC 1.1.1.27) has been investigated by calorimetric and equilibrium dialysis techniques in 0.2 M potassium phosphate buffer (pH 7.0) at various temperatures. Analysis of thermal titration curves at two temperatures (25 and 31.5 degrees) shows that the experimental enthalpy data can be rationalized assuming four independent and equivalent binding sites for the tetrameric enzyme. Binary complex formation is characterized by a negative temperature coefficient, delta cp, of the binding enthalpy, which amounts to -1300 plus or minus 53 cal/(deg mol of LDH) in the temperature range of 5-31.5 degrees. Despite the slightly smaller standard deviation resulting when polynomial regression analysis of the second degree is applied to the temperature dependence of the enthalpy values, binding enthalpies seem to be adequately represented in the temperature range studied by the equation delta H = -1.3T + 2.3, kcal/mol of LDH, T referring to the temperature in degrees C. By combination of the results obtained from equilibrium dialysis and calorimetric studies a set of apparent thermodynamic parameters for binding of NADH to LDH in 0.2 M potassium phosphate buffer at pH 7 has been established. |
