μMap-Red: Proximity Labeling by Red Light Photocatalysis

Abstract

Modern proximity labeling techniques have enabled significant advances in understanding biomolecular interactions. However, current tools primarily utilize activation modes that are incompatible with complex biological environments, limiting our ability to interrogate cell- and tissue-level microenvironments in animal models. Here, we report μMap-Red, a proximity labeling platform that uses a red-light-excited SnIV chlorin e6 catalyst to activate a phenyl azide biotin probe. We validate μMap-Red by demonstrating photonically controlled protein labeling in vitro through several layers of tissue, and we then apply our platform in cellulo to label EGFR microenvironments and validate performance with STED microscopy and quantitative proteomics. Finally, to demonstrate labeling in a complex biological sample, we deploy μMap-Red in whole mouse blood to profile erythrocyte cell-surface proteins. This work represents a significant methodological advance toward light-based proximity labeling in complex tissue environments and animal models.

The forgotten reagent of photoredox catalysis

“…4c, i) to reduce aryl azides. 68 The aminyl radical intermediates could then be used for proximity-based protein labelling in aqueous buffers. Superior media penetration of the low energy irradiation enabled erythrocyte labelling in whole blood.…”

A genetically encoded photo-proximity labeling approach for mapping protein territories

“…The recent introduction of the photo-proximity labeling (PPL) concept offers an attractive alternative to existing PL strategies. 11,12 This approach relies on the delivery of a chemical photocatalyst to a specific cellular site, e.g., through conjugation to an antibody. Selective targeting of the photocatalyst allows for the localized activation of the PL probe upon light irradiation, thereby tagging proximal proteins.…”

Amino acids with fluorescent tetrazine ethers as bioorthogonal handles for peptide modification

“…The excitation wavelength of 4b (l Exc ¼ 531 nm) ensured minimal absorption overlap with endogenous chromophores and the absence of major scattering effects, both events known to interfere with uorophores excited at shorter wavelengths (<500 nm) when used in a biological context. 36,37 We hypothesized that this advantage could counterbalance the moderate uorescence quantum yield displayed by 4b. We concluded that 4b presents the necessary trade-off between stability and reactivity to be used in vivo.…”