Three-Dimensional Structure of an Intact Glycoside Hydrolase Family 15 Glucoamylase from Hypocrea jecorina

Abstract

The three-dimensional structure of a complete Hypocrea jecorina glucoamylase has been determined at 1.8 A resolution. The presented structure model includes the catalytic and starch binding domains and traces the course of the 37-residue linker segment. While the structures of other fungal and yeast glucoamylase catalytic and starch binding domains have been determined separately, this is the first intact structure that allows visualization of the juxtaposition of the starch binding domain relative to the catalytic domain. The detailed interactions we see between the catalytic and starch binding domains are confirmed in a second independent structure determination of the enzyme in a second crystal form. This second structure model exhibits an identical conformation compared to the first structure model, which suggests that the H. jecorina glucoamylase structure we report is independent of crystal lattice contact restraints and represents the three-dimensional structure found in solution. The proposed starch binding regions for the starch binding domain are aligned with the catalytic domain in the three-dimensional structure in a manner that 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.

Purification and biochemical characterization of a novel α-glucosidase from Aspergillus niveus

“…The a-glucosidase was obtained in Khanna medium with 72 h of cultivation. The activity assay was carried out as in ''Materials and methods'', modifying pH and temperature in according to each experiment (Bott et al 2008) reveals that the family 15 domain of the glycosyl hydrolase is rich in a-helical structure, whereas the CDM domain is comprised exclusively of beta-sheet. Analysis of the three-dimensional structures of the two domains reveals a total a-helical content of 31% and a total beta-sheet content of 15%.…”

Yeast glucoamylases: molecular-genetic and structural characterization

“…Thus, the fold of this yeast glucoamylase is very similar to that of the catalytic domain of the glucoamylases from Aspergillus awamori (Aleshin et al 1992) and Hypocrea jecorina (Bott et al 2008). There is no a second domain in the S. fibuligera glucoamylases.…”

Glucoamylases from Penicillium verruculosum and Myceliophthora thermophila: Analysis of differences in activity against polymeric substrates based on 3D model structures of the intact enzymes

“…Two models of each enzyme used the 2vn7.pdb structure (TrGla, glucoamylase from T. reesei, synonym Hypocrea jecorina) as a template, while the template for the third model was glucoamylase from A. awamori (1gah.pdb). The 2vn7.pdb represents the first crystallographic structure of the intact GH15 glucoamylase including the catalytic and starch binding domains (CD and SBD, respectively) as well as the 37-residue linker segment [23], while the 1gah.pdb structure contains only the CD of the A. awamori enzyme [24]. 3D models of the PvGla and MtGla CDs, obtained with different templates, looked very similar when superimposed to each other (data not shown).…”

“…3D models of the PvGla and MtGla CDs, obtained with different templates, looked very similar when superimposed to each other (data not shown). Since analysis of the TrGla crystal structure (2vn7.pdb) revealed rather strong interactions between the SBD and CD of the enzyme, these interactions affecting the catalysis [23], it would be interesting to Table 2). The amino acid sequences of PvGla and MtGla, aligned with the TrGla sequence in Fig.…”