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Characterization and crystal structure of a thermostable glycoside hydrolase family 45 1,4-β-endoglucanase from Thielavia terrestris.
- Source :
-
Enzyme & Microbial Technology . Apr2017, Vol. 99, p32-37. 6p. - Publication Year :
- 2017
-
Abstract
- 1,4-β-Endoglucanase is one of the most important biocatalysts in modern industries. Here, a glycoside hydrolase (GH) family 45 endoglucanase from thermophilic fungus Theilavia terrestris (TtCel45A) was expressed in Pichia pastoris . The recombinant protein shows optimal activity at 60 °C, pH 4–5. The enzyme exhibits extraordinary thermostability that more than 80% activity was detected after heating at 80 °C for 2.5 h. The high resolution crystal structures of apo-form enzyme and that in complex with cellobiose and cellotetraose were solved to 1.36–1.58 Å. The protein folds into two overall regions: one is a six-stranded β-barrel, and the other one consists of several extended loops. Between the two regions lies the substrate-binding channel, which is an open cleft spanning across the protein surface. A continuous substrate-binding cleft from subsite −4 to +3 were clearly identified in the complex structures. Notably, the flexible V–VI loop ( 113 Gly- 114 Gly- 115 Asp- 116 Leu- 117 Gly- 118 Ser) is found to open in the presence of −1 sugar, with D115 and L116 swung away to yield a space to accommodate the catalytic acid D122 and the 2,5 B boat conformation of −1 sugar during transition state. Collectively, we characterized the enzyme properties of P. pastoris -expressed TtCel45A and solved high-resolution crystal structures of the enzyme. These results are of great interests in industrial applications and provide new insights into the fundamental understanding of enzyme catalytic mechanism of GH45 endoglucanases. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 01410229
- Volume :
- 99
- Database :
- Academic Search Index
- Journal :
- Enzyme & Microbial Technology
- Publication Type :
- Academic Journal
- Accession number :
- 121244250
- Full Text :
- https://doi.org/10.1016/j.enzmictec.2017.01.005