期刊
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
卷 140, 期 23, 页码 7046-7051出版社
AMER CHEMICAL SOC
DOI: 10.1021/jacs.8b03074
关键词
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资金
- National Institutes of Health [DPI AG053015, R37 GM058867]
- NOMIS Foundation
- Stanford Neurosciences Institute
- National Science Foundation Graduate Research Fellowship
- Knut and Alice Wallenberg Foundation Postdoctoral Fellowship
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [R37GM058867] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE ON AGING [DP1AG053015] Funding Source: NIH RePORTER
Bioorthogonal tools enable cell-type-specific proteomics, a prerequisite to understanding biological processes in multicellular organisms. Here we report two engineered aminoacyl-tRNA synthetases for mammalian bioorthogonal labeling: a tyrosyl (ScTyr(Y43G)) and a phenylalanyl (MmPhe(T413G)) tRNA synthetase that incorporate azide-bearing noncanonical amino acids specifically into the nascent proteomes of host cells. Azide-labeled proteins are chemoselectively tagged via azide-alkyne cycloadditions with fluorophores for imaging or affinity resins for mass spectrometric characterization. Both mutant synthetases label human, hamster, and mouse cell line proteins and selectively activate their azido-bearing amino acids over 10-fold above the canonical. ScTyr(Y43G) and MmPhe(T413G) label overlapping but distinct proteomes in human cell lines, with broader proteome coverage upon their coexpression. In mice, ScTyr(Y43G) and MmPhe(T413G) label the melanoma tumor proteome and plasma secretome. This work furnishes new tools for mammalian residue-specific bioorthogonal chemistry, and enables more robust and comprehensive cell-type-specific proteomics in live mammals.
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