SNAP-tag2: faster and brighter protein labeling

SNAP-tag is a powerful tool for labeling proteins with synthetic fluorophores in bioimaging. However, its utility in live-cell applications can be constrained by its relatively slow labeling kinetics and the limited cell permeability of its substrates. Here we introduce new labeling substrates and an engineered SNAP-tag for faster labelingin vitroand in live cells. SNAP-tag2 presents a second-order rate constant with rhodamine substrates that approaches 10⁷s^-1M^-1, a 100-fold improvement over the corresponding SNAP-tag-substrate pairs. When labeled with highly fluorogenic dyes, SNAP-tag2 also shows a 5-fold increase in fluorescence brightness relative to currently used SNAP-tag. The increased labeling kinetics and brightness of SNAP-tag2 translates into a greatly improved performance in various live-cell (super-resolution) imaging applications.