8Y33

A near-infrared fluorescent protein of de novo backbone design

8Y33 の概要

• エントリーDOI: 10.2210/pdb8y33/pdb
• 分子名称: near-infrared fluorescent protein, 3-[5-[(4-ethenyl-3-methyl-5-oxidanylidene-pyrrol-2-yl)methyl]-2-[[5-[(3-ethyl-4-methyl-5-oxidanylidene-pyrrol-2-yl)methyl]-3-(3-hydroxy-3-oxopropyl)-4-methyl-1~{H}-pyrrol-2-yl]methyl]-4-methyl-1~{H}-pyrrol-3-yl]propanoic acid (2 entities in total)
• 機能のキーワード: de novo backbone design, monomerization, near-infrared fluorescent protein, de novo protein
• 由来する生物種: synthetic construct
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 21723.38

構造登録者

Hu, X.,Xu, Y. (登録日: 2024-01-28, 公開日: 2024-02-28, 最終更新日: 2024-05-01)

主引用文献

Hu, X.,Xu, Y.,Yi, J.,Wang, C.,Zhu, Z.,Yue, T.,Zhang, H.,Wang, X.,Wu, F.,Xue, L.,Bai, L.,Liu, H.,Chen, Q.
Using Protein Design and Directed Evolution to Monomerize a Bright Near-Infrared Fluorescent Protein.
Acs Synth Biol, 13:1177-1190, 2024

PubMed Abstract: The small ultrared fluorescent protein (smURFP) is a bright near-infrared (NIR) fluorescent protein (FP) that forms a dimer and binds its fluorescence chromophore, biliverdin, at its dimer interface. To engineer a monomeric NIR FP based on smURFP potentially more suitable for bioimaging, we employed protein design to extend the protein backbone with a new segment of two helices that shield the original dimer interface while covering the biliverdin binding pocket in place of the second chain in the original dimer. We experimentally characterized 13 designs and obtained a monomeric protein with a weak fluorescence. We enhanced the fluorescence of this designed protein through two rounds of directed evolution and obtained designed monomeric smURFP (DMsmURFP), a bright, stable, and monomeric NIR FP with a molecular weight of 19.6 kDa. We determined the crystal structures of DMsmURFP both in the apo state and in complex with biliverdin, which confirmed the designed structure. The use of DMsmURFP in in vivo imaging of mammalian systems was demonstrated. The backbone design-based strategy used here can also be applied to monomerize other naturally multimeric proteins with intersubunit functional sites.

PubMed: 38552148
DOI: 10.1021/acssynbio.3c00643

実験手法

X-RAY DIFFRACTION (2.9 Å)