Optimized genetic code expansion technology for time-dependent induction of adhesion GPCR-ligand engagement.

The introduction of an engineered aminoacyl-tRNA synthetase/tRNA pair enables site-specific incorporation of unnatural amino acids (uAAs) with functionalized side chains into proteins of interest. Genetic Code Expansion (GCE) via amber codon suppression confers functionalities to proteins but can also be used to temporally control the incorporation of genetically encoded elements into proteins. Here, we report an optimized GCE system (GCEXpress) for efficient and fast uAA incorporation.

Dual-Color Fluorescence Cross-Correlation Spectroscopy to Study Protein-Protein Interaction and Protein Dynamics in Live Cells.

We present a protocol and workflow to perform live cell dual-color fluorescence cross-correlation spectroscopy (FCCS) combined with Förster Resonance Energy transfer (FRET) to study membrane receptor dynamics in live cells using modern fluorescence labeling techniques. In dual-color FCCS, where the fluctuations in fluorescence intensity represent the dynamic "fingerprint" of the respective fluorescent biomolecule, we can probe co-diffusion or binding of the receptors. FRET, with its high sensitivity to molecular distances, serves as a well-known "nanoruler" to monitor intramolecular changes.