Logic-gating the HaloTag system with Conditional-Halo-ligator 'CHalo' reagents

Philipp Mauker; Carmen Zecha; Lucas Dessen-Weissenhorn; Luciano Román-Albasini; Joyce C. M. Meiring; Julia Ines Brandmeier; Daniela Beckmann; Jan P. Prohaska; Martin Reynders; Yi Louise Li; Kristina Mrug; Hannah von Schwerin; Nynke A. Vepřek; Julia F. Gaisbauer; Leif Dehmelt; Perihan Nalbant; Jennifer Zenker; Anna S. Akhmanova; Martin Kerschensteiner; Angelika Bettina Harbauer; Julia Thorn‐Seshold; Oliver Thorn‐Seshold

doi:10.1101/2025.09.23.676741

Cell BiologybioRxivSep 23, 2025

Preprint

Logic-gating the HaloTag system with Conditional-Halo-ligator 'CHalo' reagents

Philipp Mauker iD, Carmen Zecha, Lucas Dessen-Weissenhorn, Luciano Román-Albasini iD, Joyce C. M. Meiring iD, Julia Ines Brandmeier, Daniela Beckmann, Jan P. Prohaska iD, Martin Reynders iD, Yi Louise Li iD, Kristina Mrug iD, Hannah von Schwerin, Nynke A. Vepřek iD, Julia F. Gaisbauer iD, Leif Dehmelt iD, Perihan Nalbant iD, Jennifer Zenker iD, Anna S. Akhmanova iD, Martin Kerschensteiner iD, Angelika Bettina Harbauer iD, Julia Thorn‐Seshold iD, Oliver Thorn‐Seshold iD

Abstract

HaloTag proteins spontaneously ligate onto any chemical reagent featuring a chloroalkane motif (CA). We introduce the conditional CHalo motif, which ligates to HaloTag only after uncaging by light or enzymes. (1) Photo-triggered CHalo fluorogenic reagents allow spatiotemporally-specific labeling; (2) photo-triggered CHalo heterodimerisers can photocontrol protein recruitment; and (3) enzyme-triggered CHalo reagents can durably record diverse enzyme activities, and multiplexing them should allow quantitative ratiometric recording of multiple activities in parallel. CHalo thus permits manifold extensions to the HaloTag technology. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=118 SRC="FIGDIR/small/676741v1_ufig1.gif" ALT="Figure 1"> View larger version (40K): org.highwire.dtl.DTLVardef@d18cd6org.highwire.dtl.DTLVardef@175694forg.highwire.dtl.DTLVardef@154ffd5org.highwire.dtl.DTLVardef@167fec2_HPS_FORMAT_FIGEXP M_FIG C_FIG

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