Multiplexed no-wash cellular imaging using BenzoTag, an evolved self-labeling protein.

ABSTRACT: Self-labeling proteins are powerful tools for exploring biology as they enable the precise cellular localization of a synthetic molecule, often a fluorescent dye. HaloTag7 is the most popular self-labeling protein due to its broad utility, its bio-orthogonality, and the simplicity of its chloroalkane ligand. However, reaction rates of HaloTag7 with different chloroalkane-containing substrates are highly variable and rates are only very fast for rhodamine-based dyes. This is a major limitation for the HaloTag system because fast labeling rates are critical for live-cell assays. Here, we use yeast surface display to produce a HaloTag variant, BenzoTag, with improved performance with a fluorogenic benzothiadiazole dye. Molecular evolution improved conjugation kinetics and increased the signal

SUBMITTER: Lampkin BJ 

PROVIDER: S-EPMC11487927 | biostudies-literature | 2024 Oct

REPOSITORIES: biostudies-literature