Visualization of Protein‐Specific Glycation in Living Cells via Bioorthogonal Chemical Reporter

Abstract

Due to the lack of suitable chemical tools, probing the protein-specific glycation is highly challenging. Herein, we present a strategy based on glycation chemical reporter and proximity-induced FRET signal readout for visualizing protein-specific glycation in living cells. We first developed a bioorthogonal glucose analogue, 6-azido-6-deoxy-D-glucose (6AzGlc), as a novel glycation chemical reporter. Two types of DNA probes, glycation conversion probe and protein targeting probe, were designed to attach to glycation adducts and target proteins, respectively. After the protein was glycated by 6AzGlc, two DNA probes were sequentially applied to the target protein, triggering proximityinduced FRET signal readout. This strategy was successfully used to visualize glucose glycation of several proteins, including PD-L1 and integrin. More importantly, this strategy allowed us to analyze corresponding biological functions of glycated protein in the native environment.

Bioorthogonal Probes for Analyzing Non‐Enzymatic Post‐Translational Modifications

“…Site-specific analysis [34] In situ visualization [14] ChemBioChem sulfenylated Cys through a strain-promoted group transfer reaction to give stable sulfoxide products (Figure 2B). [16] With the norbornene-derived BPs (norb-yne) Chalker's group performed a whole-cell sulfenomic analysis and found 148 new protein members of the sulfenome, which helped a deep understanding of the role of sulfenylation in redox signaling and diseases associated with oxidative stress.…”

“…Fluorescent labeling by bioorthogonal reaction allows the observation of the overall protein glycation in situ , but cannot focus on particular proteins. Herein, our group has developed a strategy for visual analysis of protein‐specific glycation in living cells based on a novel BPs and the technique of fluorescence resonance energy transfer (FRET) [14] . We first exploited a bioorthogonal glucose analogue, 6‐azido‐6‐deoxy‐D‐glucose (6AzGlc), as a novel BPs to glycate proteins in living cells.…”

“…Further functional studies revealed that the glycation of PD‐L1 and integrin impacted their binding to ligands, which may contribute to the development of targeted therapeutic methods. More importantly, this strategy is versatile, readily extended to other targeting modules and BPs, paving a new way for deep understanding of nPTMs and their biological function [14] …”

“…By virtue of these advantages, BPs have attracted considerable interest and have become an excellent tool for nPTMs visualization and proteomic investigation (Figure 1). [10a,14] Herein, we review the currently available BPs used for the study of different types of nPTMs (as summarized in Table 1), clarify their corresponding working principles, and highlight how they advance our understanding of the nPTMs biological functions.…”

Proximity-driven bioorthogonal reactionsviaDNA circuits enable the accurate aptasensing of non-nucleic acid targets

“…Moreover, aptamers can undergo structural transformation in response to target binding, enabling the conversion of target recognition into a signal output. Benefiting from these unique properties, aptamers have been exploited to construct versatile aptamer sensors (aptasensors) for the detection of targets of interest, such as proteins, 15,16 small molecules, 17,18 bacteria, 19,20 and cells. 21,22 In addition, to improve the sensitivity of aptasensors, aptamers can further couple with DNA cascade circuits for signal amplification, affording the capacity for low-abundance target detection and imaging.…”