SNAP-tag2: faster and brighter protein labeling

Kühn, S.; Nasufovic, V.; Wilhelm, J.; Kompa, J.; de Lange, E. M. F.; Lin, Y.-H.; Egoldt, C.; Fischer, J.; Lennoi, A.; Tarnawski, M.; Reinstein, J.; Vlijm, R.; Hiblot, J.; Johnsson, K.

doi:10.1101/2024.08.28.610127

bioRxiv · 10.1101/2024.08.28.610127

SNAP-tag2: faster and brighter protein labeling

Kühn, S.·Nasufovic, V.·Wilhelm, J.·Kompa, J.·de Lange, E. M. F.·Lin, Y.-H.·Egoldt, C.·Fischer, J.·Lennoi, A.·Tarnawski, M.·Reinstein, J.·Vlijm, R.·Hiblot, J.·Johnsson, K.

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Abstract

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 labeling in vitro and in live cells. SNAP-tag2 presents a second-order rate constant with rhodamine substrates that approaches 107 s-1 M-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.

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Kühn, S., Nasufovic, V., Wilhelm, J., Kompa, J., de Lange, E. M. F., Lin, Y.-H., Egoldt, C., Fischer, J., Lennoi, A., Tarnawski, M., Reinstein, J., Vlijm, R., Hiblot, J., Johnsson, K.. 2024-08-29. SNAP-tag2: faster and brighter protein labeling. https://doi.org/10.1101/2024.08.28.610127

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