Rapid Mapping of Protein Interactions Using Tag‐Transfer Photocrosslinkers
| العنوان: | Rapid Mapping of Protein Interactions Using Tag‐Transfer Photocrosslinkers |
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| المؤلفون: | Jim E. Horne, Martin Walko, Antonio N. Calabrese, Mark A. Levenstein, David J. Brockwell, Nikil Kapur, Andrew J. Wilson, Sheena E. Radford |
| المصدر: | Angewandte Chemie (International Ed. in English) |
| بيانات النشر: | Wiley, 2018. |
| سنة النشر: | 2018 |
| مصطلحات موضوعية: | 0301 basic medicine, Chemical Crosslinking, diazo compounds, Peptide, protein–protein interactions, macromolecular substances, Mass spectrometry, 01 natural sciences, Mass Spectrometry, Catalysis, Protein–protein interaction, 03 medical and health sciences, chemistry.chemical_compound, Cysteine, Protein Interaction Maps, Mesylates, chemistry.chemical_classification, Molecular Structure, Photoaffinity labeling, biology, 010405 organic chemistry, Communication, technology, industry, and agriculture, Proteins, General Medicine, General Chemistry, Photochemical Processes, Combinatorial chemistry, Communications, 0104 chemical sciences, Cross-Linking Reagents, 030104 developmental biology, chemistry, Chaperone (protein), Reagent, Diazirine, biology.protein, photoaffinity labeling |
| الوصف: | 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 methanethiosulphonate (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 initially 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). |
| وصف الملف: | application/pdf |
| تدمد: | 1521-3773 1433-7851 0044-8249 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::6bd52ca48a515e563dd60ad2ee6332bb https://doi.org/10.1002/anie.201809149 |
| حقوق: | OPEN |
| رقم الأكسشن: | edsair.doi.dedup.....6bd52ca48a515e563dd60ad2ee6332bb |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 15213773 14337851 00448249 |
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