7PCA

Functional and structural characterization of redox sensitive superfolder green fluorescent protein and variants

7PCA の概要

• エントリーDOI: 10.2210/pdb7pca/pdb
• 分子名称: Green fluorescent protein, GLYCEROL, ETHANOL, ... (5 entities in total)
• 機能のキーワード: genetically encoded biosensors, x-ray crystal structure, dynamic simulation, redox regulation, plasmodium falciparum, fluorescent protein
• 由来する生物種: Aequorea victoria (Water jellyfish, Mesonema victoria)
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 27118.57

構造登録者

Fritz-Wolf, K.,Heimsch, K.C.,Schuh, A.K.,Becker, K. (登録日: 2021-08-03, 公開日: 2022-02-16, 最終更新日: 2024-10-23)

主引用文献

Heimsch, K.C.,Gertzen, C.G.W.,Schuh, A.K.,Nietzel, T.,Rahlfs, S.,Przyborski, J.M.,Gohlke, H.,Schwarzlander, M.,Becker, K.,Fritz-Wolf, K.
Structure and Function of Redox-Sensitive Superfolder Green Fluorescent Protein Variant.
Antioxid.Redox Signal., 37:1-18, 2022

PubMed Abstract: Genetically encoded green fluorescent protein (GFP)-based redox biosensors are widely used to monitor specific and dynamic redox processes in living cells. Over the last few years, various biosensors for a variety of applications were engineered and enhanced to match the organism and cellular environments, which should be investigated. In this context, the unicellular intraerythrocytic parasite , the causative agent of malaria, represents a challenge, as the small size of the organism results in weak fluorescence signals that complicate precise measurements, especially for cell compartment-specific observations. To address this, we have functionally and structurally characterized an enhanced redox biosensor superfolder roGFP2 (sfroGFP2). SfroGFP2 retains roGFP2-like behavior, yet with improved fluorescence intensity (FI) . SfroGFP2-based redox biosensors are pH insensitive in a physiological pH range and show midpoint potentials comparable with roGFP2-based redox biosensors. Using crystallography and rigidity theory, we identified the superfolding mutations as being responsible for improved structural stability of the biosensor in a redox-sensitive environment, thus explaining the improved FI . This work provides insight into the structure and function of GFP-based redox biosensors. It describes an improved redox biosensor (sfroGFP2) suitable for measuring oxidizing effects within small cells where applicability of other redox sensor variants is limited. Improved structural stability of sfroGFP2 gives rise to increased FI . Fusion to hGrx1 (human glutaredoxin-1) provides the hitherto most suitable biosensor for measuring oxidizing effects in . This sensor is of major interest for studying glutathione redox changes in small cells, as well as subcellular compartments in general. 37, 1-18.

PubMed: 35072524
DOI: 10.1089/ars.2021.0234

実験手法

X-RAY DIFFRACTION (1.05 Å)