Rabbit liver fructose 1,6-diphosphatase. Properties of the native enzyme and their modification by subtilisin

Abstract

A simple two-step procedure is described for the purification of fructose 1, 6-diphosphatase from rabbit liver. The enzyme shows activity at neutral pH, in contrast to the previously described “alkaline” fructose diphosphatase. The neutral fructose diphosphatase has a higher molecular weight, 140,000, as compared with 130,000 for the alkaline enzyme and a somewhat different amino acid composition. Most significant is the presence of one residue of tryptophan per subunit; this amino acid is not present in the alkaline fructose diphosphatase. The subunit molecular weight of the neutral enzyme is 36,000, and it contains 4 moles of COOH-terminal alanine. Digestion with subtilisin converts the neutral enzyme to one with properties resembling those of the alkaline form. The pH optimum is shifted to pH 9 and the sensitivity toward inhibition by AMP is decreased. The molecular weight of the modified enzyme is 120,000. The results suggest that the changes in catalytic properties are associated with the removal of a tryptophan-containing peptide or peptides with molecular weights totaling approximately 6000 per subunit, and that the alkaline enzyme previously studied in this and other laboratories is a modified form of the native neutral fructose diphosphatase.