4KA9

Crystal structure analysis of single amino acid deletion mutations in EGFP

4KA9 の概要

• エントリーDOI: 10.2210/pdb4ka9/pdb
• 関連するPDBエントリー: 4KAG 4KEX
• 分子名称: Green fluorescent protein, 1,2-ETHANEDIOL, 4-(2-HYDROXYETHYL)-1-PIPERAZINE ETHANESULFONIC ACID, ... (6 entities in total)
• 機能のキーワード: beta barrel, fluorescent protein, chromophore cyclisation, single amino acid deletion mutation, cyclisation
• 由来する生物種: Aequorea victoria (Jellyfish)
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 27873.11

構造登録者

Arpino, J.A.J.,Rizkallah, P.J. (登録日: 2013-04-22, 公開日: 2014-05-07, 最終更新日: 2015-11-25)

主引用文献

Arpino, J.A.,Reddington, S.C.,Halliwell, L.M.,Rizkallah, P.J.,Jones, D.D.
Random single amino acid deletion sampling unveils structural tolerance and the benefits of helical registry shift on GFP folding and structure.
Structure, 22:889-898, 2014

PubMed Abstract: Altering a protein's backbone through amino acid deletion is a common evolutionary mutational mechanism, but is generally ignored during protein engineering primarily because its effect on the folding-structure-function relationship is difficult to predict. Using directed evolution, enhanced green fluorescent protein (EGFP) was observed to tolerate residue deletion across the breadth of the protein, particularly within short and long loops, helical elements, and at the termini of strands. A variant with G4 removed from a helix (EGFP(G4Δ)) conferred significantly higher cellular fluorescence. Folding analysis revealed that EGFP(G4Δ) retained more structure upon unfolding and refolded with almost 100% efficiency but at the expense of thermodynamic stability. The EGFP(G4Δ) structure revealed that G4 deletion caused a beneficial helical registry shift resulting in a new polar interaction network, which potentially stabilizes a cis proline peptide bond and links secondary structure elements. Thus, deletion mutations and registry shifts can enhance proteins through structural rearrangements not possible by substitution mutations alone.

PubMed: 24856363
DOI: 10.1016/j.str.2014.03.014

実験手法

X-RAY DIFFRACTION (1.58 Å)