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5RGA

Crystal Structure of Kemp Eliminase HG3 with bound transition state analogue, 277K

5RGA の概要
エントリーDOI10.2210/pdb5rga/pdb
Group depositionCrystal structures of HG-series of Kemp Eliminases at Room-temperature (G_1002148)
分子名称Kemp Eliminase HG3, 6-NITROBENZOTRIAZOLE, SULFATE ION, ... (4 entities in total)
機能のキーワードhydrolase
由来する生物種Thermoascus aurantiacus
タンパク質・核酸の鎖数2
化学式量合計69491.85
構造登録者
Broom, A.,Rakotoharisoa, R.V.,Thompson, M.C.,Fraser, J.S.,Chica, R.A. (登録日: 2020-03-19, 公開日: 2020-07-22, 最終更新日: 2024-11-06)
主引用文献Broom, A.,Rakotoharisoa, R.V.,Thompson, M.C.,Zarifi, N.,Nguyen, E.,Mukhametzhanov, N.,Liu, L.,Fraser, J.S.,Chica, R.A.
Ensemble-based enzyme design can recapitulate the effects of laboratory directed evolution in silico.
Nat Commun, 11:4808-4808, 2020
Cited by
PubMed Abstract: The creation of artificial enzymes is a key objective of computational protein design. Although de novo enzymes have been successfully designed, these exhibit low catalytic efficiencies, requiring directed evolution to improve activity. Here, we use room-temperature X-ray crystallography to study changes in the conformational ensemble during evolution of the designed Kemp eliminase HG3 (k/K 146 Ms). We observe that catalytic residues are increasingly rigidified, the active site becomes better pre-organized, and its entrance is widened. Based on these observations, we engineer HG4, an efficient biocatalyst (k/K 103,000 Ms) containing key first and second-shell mutations found during evolution. HG4 structures reveal that its active site is pre-organized and rigidified for efficient catalysis. Our results show how directed evolution circumvents challenges inherent to enzyme design by shifting conformational ensembles to favor catalytically-productive sub-states, and suggest improvements to the design methodology that incorporate ensemble modeling of crystallographic data.
PubMed: 32968058
DOI: 10.1038/s41467-020-18619-x
主引用文献が同じPDBエントリー
実験手法
X-RAY DIFFRACTION (1.86 Å)
構造検証レポート
Validation report summary of 5rga
検証レポート(詳細版)ダウンロードをダウンロード

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件を2024-11-20に公開中

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