Stability studies on pig heart mitochondrial malate dehydrogenase: the effect of salts and amino acids

Abstract

The effect of different salts and amino acids on the thermal stability and quaternary conformation of pig heart mitochondrial malate dehydrogenase ( phm-MDH) in solution has been determined. The effectiveness of salts of anions in the stabilisation of phm-MDH followed the order: Citrate > SO 4 2− ≥ Tartrate > Phosphate > F −, CH 3COO −> Cl −> Br −. Anions above and including Cl − in this series were increasingly effective in stabilising phm-MDH with a rise in salt concentration from 0.05 – 2 M, whilst Br − was destabilising under similar conditions. The effect of potassium salts of acetate, chloride and bromide at a concentration of 1 M on the quaternary conformation of phm-MDH correlated also with the relative order of anion stabilisation above, with the anions higher in the series increasingly promoting the formation of the dimeric conformation of the enzyme. The cations of the corresponding salts had a relatively neutral (Cs +, K +, Na +, (CH 3) 4N +, NH 4 +) to a destabilising ((CH 3) 4N +, NH 4 +, Li +) effect on phm-MDH. Potassium ferrocyanide and potassium ferricyanide conferred complex, concentration dependent effects on the stability of phm-MDH, unlike the salts described above. Salts of amino acids were effective in the stabilisation of phm-MDH against temperature induced changes, following the order: NaGlutamatec ≅ NaAspartate > NaGlycinate > lysine · HCl > arginine · HCl. The magnitudes and trends of the effects of these salts and amino acids on the stability and quaternary structure of phm-MDH were observed to correlate well with considerations based on the Hofmeister series of anions and solvophobic concepts as they apply to the influence of co-solvents at intermediate to higher concentrations. Other, more specific effects were also evident in the stabilisation and destabilisation of phm-MDH by low concentrations of the salts, as noted most particularly in the presence of potassium ferrocyanide and potassium ferricyanide.