Molecular cloning, expression, purification, and characterization of fructose 1,6-bisphosphate aldolase from Mycobacterium tuberculosis—a novel Class II A tetramer
A new form of the class-II D-fructose 1,6-bisphosphate aldolase (EC 4.1.2.13) of Escherichia coli (Crookes' strain) was isolated from an extract of glycerol-grown bacteria. It has a higher molecular weight (approx. 80000)than previous preparations of the enzyme and closely resembles the typical class-II aldolase from yeast in size and amino acid composition. On the other hand, its kinetic behaviour is not typical of a class-II aldolase. The enzyme has no requirement for thiol compounds either for stability or activity, added K+ ions have no effect, and the optimum pH for the cleavage activity is unusually high. The class-II enzymes from the prokaryote E. coli and the eukaryote yeast show no immunological identity. However, the similarity of their structures suggests that they have evolved from a common ancestor.
Two proteins have been purified from culture filtrates of
Mycobacterium tuberculosis
, H
37
Ra strain by a procedure combining gel filtration, diethylaminoethyl (DEAE)-cellulose chromatography, and zone electrophoresis. The two proteins are similar in molecular weight but differ slightly in charge. The faster migrating protein, designated a
1
, is not antigenic. The slower migrating protein, designated a
2
, is antigenic both with respect to antisera and as a skin-testing antigen.
Purification and characterization of the class-II D-fructose 1,6-bisphosphate aldolase from Escherichia coli (Crookes' strain)