7Q3K

Computationally designed thioredoxin subjected to stability optimizing mutations.

7Q3K の概要

• エントリーDOI: 10.2210/pdb7q3k/pdb
• 分子名称: eMM9, SULFATE ION (2 entities in total)
• 機能のキーワード: de-novo protein design, deep mutational scanning, stability, screening, de novo protein
• 由来する生物種: synthetic construct
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 39252.40

構造登録者

Norrild, R.K.,Johansson, K.E.,O'Shea, C.,Lindorff-Larsen, K.,Winther, J.R.,Morth, J.P. (登録日: 2021-10-27, 公開日: 2022-11-16, 最終更新日: 2024-02-07)

主引用文献

Norrild, R.K.,Johansson, K.E.,O'Shea, C.,Morth, J.P.,Lindorff-Larsen, K.,Winther, J.R.
Increasing protein stability by inferring substitution effects from high-throughput experiments.
Cell Rep Methods, 2:100333-100333, 2022

PubMed Abstract: We apply a computational model, global multi-mutant analysis (GMMA), to inform on effects of most amino acid substitutions from a randomly mutated gene library. Using a high mutation frequency, the method can determine mutations that increase the stability of even very stable proteins for which conventional selection systems have reached their limit. As a demonstration of this, we screened a mutant library of a highly stable and computationally redesigned model protein using an genetic sensor for folding and assigned a stability effect to 374 of 912 possible single amino acid substitutions. Combining the top 9 substitutions increased the unfolding energy 47 to 69 kJ/mol in a single engineering step. Crystal structures of stabilized variants showed small perturbations in helices 1 and 2, which rendered them closer in structure to the redesign template. This case study illustrates the capability of the method, which is applicable to any screen for protein function.

PubMed: 36452862
DOI: 10.1016/j.crmeth.2022.100333

実験手法

X-RAY DIFFRACTION (2.25 Å)