期刊
BIOCHEMISTRY
卷 57, 期 31, 页码 4747-4752出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.biochem.8b00397
关键词
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资金
- Deutsche Forschungsgemeinschaft (DFG) [SPP1623, TRR83]
- EMBL Interdisciplinary Postdoc (EI3POD) program under Marie Sklodowska-Curie Actions COFUND [664726]
Mapping of weak and hence transient interactions between low-abundance interacting molecules is still a major challenge in systems biology and protein biochemistry. Therefore, additional system-wide acting tools are needed to determine protein interactomics. Most important are reagents that can be applied at any kind of protein interface and the possibility to enrich cross-linked fragments with high efficiency. In this study, we report the synthesis of a novel noncanonical amino acid that features a diazirine group for ultraviolet cross-linking as well as an alkyne group for labeling by click chemistry. This bifunctional amino acid, called PrDiAzK, may be inserted into almost any protein interface with minimal structural perturbation using genetic code expansion. We demonstrate that PrDiAzK can be site-selectively incorporated into proteins in both bacterial and mammalian cell cultures, and we show that PrDiAzK allows protein labeling as well as cross-linking. In addition, we tested PrDiAzK for proteome-wide incorporation via stochastic orthogonal recoding of translation, implying potential applications in system-wide mapping of protein-protein interactions in the future.
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