Thermostabilization of a thermophilic 1,4-α-glucan branching enzyme through C-terminal truncation.

Thermophilic proteins are useful for the detailed investigation of thermostability because they function efficiently at high temperatures. Comparison of the amino acid sequences and three-dimensional structures of mesophilic and thermophilic 1,4-α-glucan branching enzymes (GBEs) shows that the amino acid sequence of the last 26 residues at the C-terminal end of the GBE from Geobacillus thermoglucosidans STB02 (GBEGt , GenBank accession no. KJ660983) are not conserved, and that their 3-dimensional structure is flexible. These residues appear to be modified based upon a balance between flexibility and rigidity that is related to thermostability. In this study, a truncated mutant of GBEGt made by removing the last 26 residues from its C-terminal end was found to have increased thermostability and solubility, compared with the wild-type enzyme. Additionally, truncation of a portion of the C-terminus resulted in a decrease in aqueous stability. The circular dichroism spectra of GBEGt and GBEGt ΔC were also found to be different. These results suggest that deletion of flexible residues at the C-terminal end of GBEGt , which are located on the surface of the enzyme, enhances the thermostability of the enzyme without significantly compromising its enzymatic activity.