THE CRYSTAL-STRUCTURE OF A CLASS-II FRUCTOSE-1,6-BISPHOSPHATE ALDOLASE SHOWS A NOVEL BINUCLEAR METAL-BINDING ACTIVE-SITE EMBEDDED IN A FAMILIAR FOLD

Background: Aldolases catalyze a variety of condensation and cleavage reactions, with exquisite control on the stereochemistry. These enzyme s, therefore, are attractive catalysts for synthetic chemistry. There are two classes of aldolase: class I aldolases utilize Schiff base for mation with an active-site lysine whilst class II enzymes require a di valent metal ion, in particular zinc. Fructose-1,6-bisphosphate aldola se (FBP-aldolase) is used in gluconeogenesis and glycolysis; the enzym e controls the condensation of dihydroxyacetone phosphate with glycera ldehyde-3-phosphate to yield fructose-1,6-bisphosphate, Structures are available for class I FBP-aldolases but there is a paucity of detail on the class II enzymes. Characterization is sought to enable a dissec tion of structure/activity relationships which may assist the construc tion of designed aldolases for use as biocatalysts in synthetic chemis try. Results: The structure of the dimeric class II FBP-aldolase from Escherichia coli has been determined using data to 2.5 Angstrom resolu tion. The asymmetric unit is one subunit which presents a familiar fol d, the (alpha/beta)(8) barrel. The active centre, at the C-terminal en d of the barrel, contains a novel bimetallic-binding site with two met al ions 6.2 Angstrom apart. One ion, the identity of which is not cert ain, is buried and may play a structural or activating role. The other metal ion is zinc and is positioned at the surface of the barrel to p articipate in catalysis. Conclusions: Comparison of the structure with a class II fuculose aldolase suggests that these enzymes may share a common mechanism. Nevertheless, the class II enzymes should be subdivi ded into two categories on consideration of subunit size and fold, qua ternary structure and metal-ion binding sites. (C) Current Biology Ltd