3WP5

The crystal structure of mutant CDBFV E109A from Neocallimastix patriciarum

3WP5 の概要

• エントリーDOI: 10.2210/pdb3wp5/pdb
• 関連するPDBエントリー: 3WP4 3WP6
• 分子名称: CDBFV (2 entities in total)
• 機能のキーワード: xylanase, regulatory n-terminal region, disulfide bond, industrial enzyme, beta-jellyroll fold, hydrolase
• 由来する生物種: Neocallimastix patriciarum
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 24635.00

構造登録者

Cheng, Y.S.,Chen, C.C.,Huang, C.H.,Huang, T.Y.,Ko, T.P.,Huang, J.W.,Wu, T.H.,Liu, J.R.,Guo, R.T. (登録日: 2014-01-09, 公開日: 2014-03-19, 最終更新日: 2023-11-08)

主引用文献

Cheng, Y.S.,Chen, C.C.,Huang, C.H.,Ko, T.P.,Luo, W.,Huang, J.W.,Liu, J.R.,Guo, R.T.
Structural analysis of a glycoside hydrolase family 11 xylanase from Neocallimastix patriciarum: insights into the molecular basis of a thermophilic enzyme.
J.Biol.Chem., 289:11020-11028, 2014

PubMed Abstract: The catalytic domain of XynCDBFV, a glycoside hydrolase family 11 (GH11) xylanase from ruminal fungus Neocallimastix patriciarum previously engineered to exhibit higher specific activity and broader pH adaptability, holds great potential in commercial applications. Here, the crystal structures of XynCDBFV and its complex with substrate were determined to 1.27-1.43 Å resolution. These structures revealed a typical GH11 β-jelly-roll fold and detailed interaction networks between the enzyme and ligands. Notably, an extended N-terminal region (NTR) consisting of 11 amino acids was identified in the XynCDBFV structure, which is found unique among GH11 xylanases. The NTR is attached to the catalytic core by hydrogen bonds and stacking forces along with a disulfide bond between Cys-4 and Cys-172. Interestingly, the NTR deletion mutant retained 61.5% and 19.5% enzymatic activity at 55 °C and 75 °C, respectively, compared with the wild-type enzyme, whereas the C4A/C172A mutant showed 86.8% and 23.3% activity. These results suggest that NTR plays a role in XynCDBFV thermostability, and the Cys-4/Cys-172 disulfide bond is critical to the NTR-mediated interactions. Furthermore, we also demonstrated that Pichia pastoris produces XynCDBFV with higher catalytic activity at higher temperature than Escherichia coli, in which incorrect NTR folding and inefficient disulfide bond formation might have occurred. In conclusion, these structural and functional analyses of the industrially favored XynCDBFV provide a molecular basis of NTR contribution to its thermostability.

PubMed: 24619408
DOI: 10.1074/jbc.M114.550905

実験手法

X-RAY DIFFRACTION (1.32 Å)