9PUB

Biotin-streptavidin binding

9PUB の概要

• エントリーDOI: 10.2210/pdb9pub/pdb
• 関連するPDBエントリー: 9PUA
• 分子名称: Streptavidin, BIOTIN, GLYCEROL, ... (4 entities in total)
• 機能のキーワード: protein binding
• 由来する生物種: Streptomyces avidinii
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 27143.39

構造登録者

Whitby, F.G. (登録日: 2025-07-30, 公開日: 2025-09-03, 最終更新日: 2026-03-25)

主引用文献

Giesler, R.J.,Woodham, P.C.S.,Draper, S.R.E.,Spaltenstein, P.,Whitby, F.G.,Hill, C.P.,Kay, M.S.
Protein chirality as a determinant of ligand affinity: insights from l- and d-streptavidin.
Chem Sci, 16:23342-23350, 2025

PubMed Abstract: Streptavidin enjoys numerous biotechnological applications due to its extraordinarily high affinity for biotin and the ease of biotinylation of targets through chemical or biological means. However, two main drawbacks limit its use in therapeutic and diagnostic applications: high immunogenicity and endogenous biotin interference. We propose that a mirror-image biotin/streptavidin system could solve these problems due to the minimal immunogenicity of mirror-image (d-) proteins and the expected lower binding affinity between non-natural d-streptavidin and natural d-biotin. To comprehensively address this problem, we first synthesized the l- and d-enantiomers of streptavidin using a three-segment native chemical ligation approach. This synthesis was enabled by temporarily solubilizing an aggregation-prone peptide segment with a Glu-based AlHx 'helping hand' linker. We developed a novel high-efficiency folding protocol and characterized the synthetic proteins by circular dichroism, size-exclusion chromatography, and binding to natural d-(+)-biotin and the mirror-image l-(-)-biotin isothermal titration calorimetry. We found a 200-million-fold difference in affinity between streptavidin and its matched mismatched biotin enantiomers that renders these systems functionally orthogonal. To gain further insight into how (-)-biotin binds recombinant streptavidin, we solved high-resolution X-ray crystal structures for both the matched and mismatched interactions. This work demonstrates the high degree of stereospecificity of the streptavidin/biotin interaction and the potential utility of a mirror-image biotin/streptavidin system for therapeutic and diagnostic applications.

PubMed: 41210276
DOI: 10.1039/d5sc06380a

実験手法

X-RAY DIFFRACTION (0.953 Å)