Purification and characterization of aldolase from the parasitic protozoan Eimeria stiedai (Coccidia)
1. 1. Fructose-bisphosphate aldolase from unsporulated oocysts of Eimeria stiedai has been purified to homogeneity and identified as a class I aldolase. 2. 2. The enzyme had a molecular weight of 160,000 and a subunit molecular weight of 40,000. 3. 3. The specific activity of purified preparations with fructose 1,6-bisphosphate as substrate was 12 units/mg, which corresponded to a molecular activity of 1920 units/micromole. The enzyme was also active with fructose 1-phosphate but at one-sixteenth the rate with fructose 1,6-bisphosphate. 4. 4. The Michaelis constants at pH 7.5 were 5–9 × 10?6 M and 2.6 × 10?2 M for fructose 1,6-bisphosphate and fructose 1-phosphate, respectively. 5. 5. Like other class I aldolases, it was inactivated by treatment with carboxypeptidase A and pyridoxal phosphate. 6. 6. Aldolase from E. stiedai had a broad pH optimum from 6.5 to 8.5 and it was more active on the acidic side of the pH optimum than rabbit muscle aldolase. The difference was apparently due to tighter binding of the C-6 phosphate of fructose 1,6-bisphosphate to E. stiedai aldolase than to rabbit muscle aldolase since, at pH 5.5, the K1 for fructose 6-phosphate was lower and the degree of inactivation by glyceralde 3-phosphate was higher with the former enzyme than the latter.