5WJ4

Crystal structure of redox-sensitive green fluorescent protein Clover mutant roClover1

5WJ4 の概要

• エントリーDOI: 10.2210/pdb5wj4/pdb
• 分子名称: Green fluorescent protein (2 entities in total)
• 機能のキーワード: fluorescent protein, clover, gfp, beta barrel
• 由来する生物種: Aequorea victoria (Jellyfish)
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 53868.77

構造登録者

Liu, C.,Campbell, B.C.,Petsko, G.A. (登録日: 2017-07-21, 公開日: 2018-01-17, 最終更新日: 2026-03-18)

主引用文献

Campbell, B.C.,Petsko, G.A.,Liu, C.F.
Crystal Structure of Green Fluorescent Protein Clover and Design of Clover-Based Redox Sensors.
Structure, 26:225-237.e3, 2018

PubMed Abstract: We have determined the crystal structure of Clover, one of the brightest fluorescent proteins, and found that its T203H/S65G mutations relative to wild-type GFP lock the critical E222 side chain in a fixed configuration that mimics the major conformer of that in EGFP. The resulting equilibrium shift to the predominantly deprotonated chromophore increases the extinction coefficient (EC), opposes photoactivation, and is responsible for the bathochromic shift. Clover's brightness can further be attributed to a π-π stacking interaction between H203 and the chromophore. Consistent with these observations, the Clover G65S mutant reversed the equilibrium shift, dramatically decreased the EC, and made Clover photoactivatable under conditions that activated photoactivatable GFP. Using the Clover structure, we rationally engineered a non-photoactivatable redox sensor, roClover1, and determined its structure as well as that of its parental template, roClover0.1. These high-resolution structures provide deeper insights into structure-function relationships in GFPs and may aid the development of excitation-improved ratiometric biosensors.

PubMed: 29307487
DOI: 10.1016/j.str.2017.12.006

実験手法

X-RAY DIFFRACTION (1.631 Å)