10-02: The three-dimensional structure of an intact glucoamylase gives insight on how substrate is directed towards the active site

We present the three-dimensional structure of Hypocrea jecorina glucoamylase at 1.8 Å resolution. The structure model includes both the catalytic domain and the starch binding domain as well as the glycosylated linker segment between these domains. This is the first intact structure of a glucoamylase. Previously, structure models of only catalytic or starch-binding domains have been available and these have been of other fungal and yeast glucoamylases. That the protein is intact in the model allows visualization of the juxtaposition of the starch-binding domain relative to the catalytic domain. One of the proposed starch binding regions on the starch-binding domain are in close proximity of the active site on the catalytic domain. This supports the hypothesis that the starch-binding domain serves to target the glucoamylase at sites where the starch granular matrix is disrupted and where the enzyme might most effectively function. The detailed interactions between the catalytic and the starch-binding domains are confirmed by two independent structure determinations of the enzyme in two different crystal forms. The two structure models exhibit an identical conformation and show the same positioning of the starch-binding domain relative to the CD. This, in turn, suggests that the H. jecorina glucoamylase structure we present not only is independent of crystal lattice contacts but that it also represents the three-dimensional structure found in solution.