4MLI

Crystal structure of the SpyTag/SpyCatcher complex

4MLI の概要

• エントリーDOI: 10.2210/pdb4mli/pdb
• 関連するPDBエントリー: 4MLS
• 分子名称: Fibronectin binding protein, SpyTag (3 entities in total)
• 機能のキーワード: isopeptide bond, spycatcher, protein engineering, peptide binding protein
• 由来する生物種: Streptococcus pyogenes; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 27782.62

構造登録者

Li, L.,Fierer, J.O.,Rapoport, T.A.,Howarth, M. (登録日: 2013-09-06, 公開日: 2013-11-13, 最終更新日: 2024-10-16)

主引用文献

Li, L.,Fierer, J.O.,Rapoport, T.A.,Howarth, M.
Structural Analysis and Optimization of the Covalent Association between SpyCatcher and a Peptide Tag.
J.Mol.Biol., 426:309-317, 2014

PubMed Abstract: Peptide tagging is a key strategy for observing and isolating proteins. However, the interactions of proteins with peptides are nearly all rapidly reversible. Proteins tagged with the peptide SpyTag form an irreversible covalent bond to the SpyCatcher protein via a spontaneous isopeptide linkage, thereby offering a genetically encoded way to create peptide interactions that resist force and harsh conditions. Here, we determined the crystal structure of the reconstituted covalent complex of SpyTag and SpyCatcher at 2.1Å resolution. The structure showed the expected reformation of the β-sandwich domain seen in the parental streptococcal adhesin, but flanking sequences at both N- and C-termini of SpyCatcher were disordered. In addition, only 10 out of 13 amino acids of the SpyTag peptide were observed to interact with SpyCatcher, pointing to specific contacts important for rapid split protein reconstitution. Based on these structural insights, we expressed a range of SpyCatcher variants and identified a minimized SpyCatcher, 32 residues shorter, that maintained rapid reaction with SpyTag. Together, these results give insight into split protein β-strand complementation and enhance a distinct approach to ultrastable molecular interaction.

PubMed: 24161952
DOI: 10.1016/j.jmb.2013.10.021

実験手法

X-RAY DIFFRACTION (2.1 Å)