7EPJ

Crystal structure of E.coli CcdB mutant V46L

7EPJ の概要

• エントリーDOI: 10.2210/pdb7epj/pdb
• 分子名称: Toxin CcdB, CHLORIDE ION (3 entities in total)
• 機能のキーワード: ccdb, topoisomerase poison, global suppressor, toxin
• 由来する生物種: Escherichia coli K-12
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 11806.44

構造登録者

Manjunath, K.,Goyal, P.,Varadarajan, R. (登録日: 2021-04-26, 公開日: 2022-06-08, 最終更新日: 2023-11-29)

主引用文献

Chattopadhyay, G.,Bhowmick, J.,Manjunath, K.,Ahmed, S.,Goyal, P.,Varadarajan, R.
Mechanistic insights into global suppressors of protein folding defects.
Plos Genet., 18:e1010334-e1010334, 2022

PubMed Abstract: Most amino acid substitutions in a protein either lead to partial loss-of-function or are near neutral. Several studies have shown the existence of second-site mutations that can rescue defects caused by diverse loss-of-function mutations. Such global suppressor mutations are key drivers of protein evolution. However, the mechanisms responsible for such suppression remain poorly understood. To address this, we characterized multiple suppressor mutations both in isolation and in combination with inactive mutants. We examined six global suppressors of the bacterial toxin CcdB, the known M182T global suppressor of TEM-1 β-lactamase, the N239Y global suppressor of p53-DBD and three suppressors of the SARS-CoV-2 spike Receptor Binding Domain. When coupled to inactive mutants, they promote increased in-vivo solubilities as well as regain-of-function phenotypes. In the case of CcdB, where novel suppressors were isolated, we determined the crystal structures of three such suppressors to obtain insight into the specific molecular interactions responsible for the observed effects. While most individual suppressors result in small stability enhancements relative to wildtype, which can be combined to yield significant stability increments, thermodynamic stabilisation is neither necessary nor sufficient for suppressor action. Instead, in diverse systems, we observe that individual global suppressors greatly enhance the foldability of buried site mutants, primarily through increase in refolding rate parameters measured in vitro. In the crowded intracellular environment, mutations that slow down folding likely facilitate off-pathway aggregation. We suggest that suppressor mutations that accelerate refolding can counteract this, enhancing the yield of properly folded, functional protein in vivo.

PubMed: 36037221
DOI: 10.1371/journal.pgen.1010334

実験手法

X-RAY DIFFRACTION (1.354 Å)