Chemical Synthesis of Diubiquitin‐Based Photoaffinity Probes for Selectively Profiling Ubiquitin‐Binding Proteins

Liang, Jun; Zhang, Lin; Tan, Xiang-Long; Qi, Yun‐Kun; Feng, Shan; Deng, Haiteng; Yan, YiJing; Zheng, Ji‐Min; Liu, Lei; Tian, Changlin

doi:10.1002/anie.201611659

Cited by 40 publications

(28 citation statements)

References 56 publications

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“…Photocrosslinkers such as diazirines can address this challenge and have been used previously to tag Cys residues in peptides . Diazirine‐generated carbenes react with proteins in ns, yet long irradiation times (minutes to hours) are often required to generate acceptable crosslink yields due to the use of low intensity lamps.…”

Section: Figuresupporting

confidence: 89%

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Rapid Mapping of Protein Interactions Using Tag‐Transfer Photocrosslinkers

et al. 2018

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Analysing protein complexes by chemical crosslinking‐mass spectrometry (XL‐MS) is limited by the side‐chain reactivities and sizes of available crosslinkers, their slow reaction rates, and difficulties in crosslink enrichment, especially for rare, transient or dynamic complexes. Here we describe two new XL reagents that incorporate a methanethiosulfonate (MTS) group to label a reactive cysteine introduced into the bait protein, and a residue‐unbiased diazirine‐based photoactivatable XL group to trap its interacting partner(s). Reductive removal of the bait transfers a thiol‐containing fragment of the crosslinking reagent onto the target that can be alkylated and located by MS sequencing and exploited for enrichment, enabling the detection of low abundance crosslinks. Using these reagents and a bespoke UV LED irradiation platform, we show that maximum crosslinking yield is achieved within 10 seconds. The utility of this “tag and transfer” approach is demonstrated using a well‐defined peptide/protein regulatory interaction (BID80‐102/MCL‐1), and the dynamic interaction interface of a chaperone/substrate complex (Skp/OmpA).

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Section: Figuresupporting

confidence: 89%

“…By contrast, a 6 W Hg‐Xe lamp achieved maximal yields after ca. 20 minutes with heating of 10 °C (Figure a,b, Figure S4c), consistent with previous reports ,. The UV LED design thus improves the reaction rate 130‐fold, and decreases heating 6‐fold.…”

Section: Figurementioning

confidence: 99%

Rapid Mapping of Protein Interactions Using Tag‐Transfer Photocrosslinkers

et al. 2018

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“…The 3‐methyl‐3 H ‐diazirin‐3‐yl‐containing linker 5 , on the other hand, afforded six labeled oridonin isomers in a much higher labeling efficiency of 69 %, although two of them are one oridonin labeled with two molecules of linker 5 . To further improve the labeling efficiency of linker 5, a volatile 3‐methyl‐3 H ‐diazirin‐3‐yl‐containing linker, 3‐(2‐azidoethyl)‐3‐methyl‐3 H ‐diazirine (linker 6 ) was designed and synthesized. As shown in Figure and Figure S1, after the completion of the reaction, unreacted volatile linkers 6 and/or by‐products of linker 6 were readily removed under vacuum as indicated by 1 H NMR analysis.…”

Section: Resultsmentioning

confidence: 99%

Functionality‐Independent DNA Encoding of Complex Natural Products

et al. 2019

Angewandte Chemie

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DNA encoded chemical libraries (DELs) link the powers of genetics and chemical synthesis via combinatorial optimization. Through combinatorial chemistry, DELs can grow to the unprecedented size of billions to trillions. To take full advantage of the DEL approach, linking the power of genetics directly to chemical structures would offer even greater diversity in a finite chemical world. Natural products have evolved an incredible structural diversity along with their biological evolution. Herein, we used traditional Chinese medicines (TCMs) as examples in a late‐stage modification toolbox approach to annotate these complex organic compounds with amplifiable DNA barcodes, which could be easily incorporated into a DEL. The method of end‐products labeling also generates a cluster of isomers with a single DNA tag at different sites. These isomers provide an additional spatial diversity for multiple accessible pockets of targeted proteins. Notably, a novel PARP1 inhibitor from TCM has been identified from the natural products enriched DEL (nDEL).

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“…Conventional photoactivatable groups include benzophenone, aryl azide, and diazirine. Upon photoirradiation, these photoactive groups generate highly reactive species that react with adjacent species, resulting in direct cross‐linking of the TP with complexed species –. Thus, fluorogenic PAL has gained increasing attention by directly reporting the labeling efficiency under in vitro or cellular conditions.…”

Section: Fluorogenic Bifunctional Linkersmentioning

confidence: 99%

Emerging Applications of Fluorogenic and Non‐fluorogenic Bifunctional Linkers

Liu

2018

Chemistry A European J

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Homo- and hetero-bifunctional linkers play vital roles in constructing a variety of functional systems, ranging from protein bioconjugates with drugs and functional agents, to surface modification of nanoparticles and living cells, and to the cyclization/dimerization of synthetic polymers and biomolecules. Conventional approaches for assaying conjugation extents typically rely on ex situ techniques, such as mass spectrometry, gel electrophoresis, and size-exclusion chromatography. If the conjugation process involving bifunctional linkers was rendered fluorogenic, then in situ monitoring, quantification, and optical tracking/visualization of relevant processes would be achieved. In this review, conventional non-fluorogenic linkers are first discussed. Then the focus is on the evolution and emerging applications of fluorogenic bifunctional linkers, which are categorized into hetero-bifunctional single-caging fluorogenic linkers, homo-bifunctional double-caging fluorogenic linkers, and hetero-bifunctional double-caging fluorogenic linkers. In addition, stimuli-cleavable bifunctional linkers designed for both conjugation and subsequent site-specific triggered release are also summarized.

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Chemical Synthesis of Diubiquitin‐Based Photoaffinity Probes for Selectively Profiling Ubiquitin‐Binding Proteins

Cited by 40 publications

References 56 publications

Rapid Mapping of Protein Interactions Using Tag‐Transfer Photocrosslinkers

Rapid Mapping of Protein Interactions Using Tag‐Transfer Photocrosslinkers

Functionality‐Independent DNA Encoding of Complex Natural Products

Emerging Applications of Fluorogenic and Non‐fluorogenic Bifunctional Linkers

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