Optimization of the genetic code expansion technology for intracellular labelling and single-molecule tracking of proteins in genomically re-coded <i>E. coli</i>.

ABSTRACT: Single-molecule tracking (SMT) is a powerful tool for real-time studies of protein interactions in living cells. Dye-labelled SNAP-tag and HaloTag self-labelling proteins have simplified SMT significantly, due to their superior photophysical properties compared to fluorescent proteins. However, due to their size, fusion of these tags to a protein of interest often results in loss of protein function. We introduce FLORENCE - a universal labelling method for SMT, based on genetic code expansion (GCE). We overcome significant caveats related to re-coded strains, vectors, and dyes and report successful tracking of site-specifically intracellularly labelled proteins in genomically re-coded E. coli. Our findings establish a robust in vivo protein-labelling strategy, expanding the c

SUBMITTER: Ilievski F 

PROVIDER: S-EPMC12684252 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature