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| Item Type: | Article |
|---|---|
| Title: | Structure and function of the Bacillus hybrid enzyme GluXyn-1: Native-like jellyroll fold preserved after insertion of autonomous globular domain |
| Creators Name: | Ay, J., Goetz, F., Borriss, R. and Heinemann, U. |
| Abstract: | The 1,3-1,4-beta-glucanase from Bacillus macerans (wtGLU) and the 1, 4-beta-xylanase from Bacillus subtilis (wtXYN) are both single-domain jellyroll proteins catalyzing similar enzymatic reactions. In the fusion protein GluXyn-1, the two proteins are joined by insertion of the entire XYN domain into a surface loop of cpMAC-57, a circularly permuted variant of wtGLU. GluXyn-1 was generated by protein engineering methods, produced in Escherichia coli and shown to fold spontaneously and have both enzymatic activities at wild-type level. The crystal structure of GluXyn-1 was determined at 2.1 A resolution and refined to R = 17.7% and R(free) = 22.4%. It shows nearly ideal, native-like folding of both protein domains and a small, but significant hinge bending between the domains. The active sites are independent and accessible explaining the observed enzymatic activity. Because in GluXyn-1 the complete XYN domain is inserted into the compact folding unit of GLU, the wild-type-like activity and tertiary structure of the latter proves that the folding process of GLU does not depend on intramolecular interactions that are short-ranged in the sequence. Insertion fusions of the GluXyn-1 type may prove to be an easy route toward more stable bifunctional proteins in which the two parts are more closely associated than in linear end-to-end protein fusions. |
| Keywords: | Bacillus, Endo-1,4-beta Xylanases, Glycoside Hydrolases, Plasmids, Protein Folding, Recombinant Fusion Proteins, Structure-Activity Relationship, Xylosidases |
| Source: | Proceedings of the National Academy of Sciences of the United States of America |
| ISSN: | 0027-8424 |
| Publisher: | National Academy of Sciences |
| Volume: | 95 |
| Number: | 12 |
| Page Range: | 6613-6618 |
| Date: | 9 June 1998 |
| Official Publication: | http://www.pnas.org/content/95/12/6613.abstract |
| PubMed: | View item in PubMed |
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