One-pot dual protein labeling for simultaneous mechanical and fluorescent readouts in optical tweezers.

ABSTRACT: Optical tweezers are widely used in the study of biological macromolecules but are limited by their one-directional probing capability, potentially missing critical conformational changes. Combining fluorescence microscopy with optical tweezers, employing Förster resonance energy transfer (FRET) pairs, addresses this issue. When integrating fluorescence microscopy with optical tweezers, orthogonal protein conjugation methods are needed to enable simultaneous, site-specific attachment of fluorophores and DNA handles, commonly used to apply force to molecules of interest. In this study, we utilized commercially available reagents for dual site-specific labeling of the homodimeric heat shock protein 90 (Hsp90) using thiol-maleimide and inverse electron demand Diels-Alder cycloaddition (IEDDAC

SUBMITTER: Silbermann LM 

PROVIDER: S-EPMC11915586 | biostudies-literature | 2025 Apr

REPOSITORIES: biostudies-literature