Stabilization of phosphofructokinase during air-drying with sugars and sugar/transition metal mixtures

Abstract

Phosphofructokinase (PFK) purified from rabbit skeletal muscle is fully inactivated after air-drying and rehydration. The addition of trehalose, maltose, or sucrose to the enzyme solution prior to rapid drying results in a recovery of almost 70% of the original activity, whereas about 30% is recovered during slow drying. Similar stabilization is seen with up to 200 mM lactose, but at higher concentrations the sugar comes out of solution during drying, and there is a dramatic drop in the activity recovered. Glucose at concentrations up to 500 mM is ineffective at protecting air-dried PFK. Addition of ionic zinc to enzyme-sugar mixtures prior to drying greatly enhances the stabilization imparted by the above sugars, but zinc alone affords no protection. Several other organic solutes (proline, glycine, trimethylamine N-oxide, glycerol, and myo-inositol) that afford cryoprotection to PFK, an effect enhanced by the addition of zinc, do not stabilize the enzyme during air-drying, even if zinc is present. The addition of ionic copper, cobalt, or nickel to trehalose-PFK solution prior to rapid drying results in a large increase in the activity recovered, and the presence of cadmium or manganese leads to a minor increase. Magnesium and calcium are ineffective in this respect. During slow drying, the presence of cadmium or calcium leads to increased preservation, magnesium and manganese have no influence on stabilization, and copper and nickel inactive the enzyme.