Single-Chain Lanthanide Luminescence Biosensors for Cell-Based Imaging and Screening of Protein-Protein Interactions

Ting Chen; Ha Pham; Ali Mohamadi; Lawrence W Miller

doi:10.1016/j.isci.2020.101533

Single-Chain Lanthanide Luminescence Biosensors for Cell-Based Imaging and Screening of Protein-Protein Interactions

iScience. 2020 Sep 6;23(9):101533. doi: 10.1016/j.isci.2020.101533. eCollection 2020 Sep 25.

Authors

Ting Chen¹, Ha Pham¹, Ali Mohamadi¹, Lawrence W Miller¹

Affiliation

¹ Department of Chemistry, University of Illinois at Chicago, Chicago, IL, USA.

Abstract

Lanthanide-based, Förster resonance energy transfer (LRET) biosensors enabled sensitive, time-gated luminescence (TGL) imaging or multiwell plate analysis of protein-protein interactions (PPIs) in living cells. We prepared stable cell lines that expressed polypeptides composed of an alpha helical linker flanked by a Tb(III) complex-binding domain, GFP, and two interacting domains at each terminus. The PPIs examined included those between FKBP12 and the rapamycin-binding domain of m-Tor (FRB) and between p53 (1-92) and HDM2 (1-128). TGL microscopy revealed dramatic differences (>500%) in donor- or acceptor-denominated, Tb(III)-to-GFP LRET ratios between open (unbound) and closed (bound) states of the biosensors. We observed much larger signal changes (>2,500%) and Z'-factors of 0.5 or more when we grew cells in 96- or 384-well plates and analyzed PPI changes using a TGL plate reader. The modular design and exceptional dynamic range of lanthanide-based LRET biosensors will facilitate versatile imaging and cell-based screening of PPIs.

Keywords: Biomolecular Engineering; Molecular Interaction; Molecular Spectroscopy Techniques; Sensor.

Grants and funding

R01 GM081030/GM/NIGMS NIH HHS/United States