Labeling, detection and identification of newly synthesized proteomes with bioorthogonal non-canonical amino-acid tagging

@article{Dieterich2007LabelingDA,
  title={Labeling, detection and identification of newly synthesized proteomes with bioorthogonal non-canonical amino-acid tagging},
  author={Daniela C. Dieterich and Jennifer Lee and A. James Link and Johannes Graumann and David A. Tirrell and Erin Margaret Schuman},
  journal={Nature Protocols},
  year={2007},
  volume={2},
  pages={532-540}
}
A major aim of proteomics is the identification of proteins in a given proteome at a given metabolic state. This protocol describes the step-by-step labeling, purification and detection of newly synthesized proteins in mammalian cells using the non-canonical amino acid azidohomoalanine (AHA). In this method, metabolic labeling of newly synthesized proteins with AHA endows them with the unique chemical functionality of the azide group. In the subsequent click chemistry tagging reaction, azide… 
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252 Citations

Metabolic labeling with noncanonical amino acids and visualization by chemoselective fluorescent tagging.

A fluorescence-based method to follow proteome-wide patterns of newly synthesized proteins in cultured cells, tissue slices, and a whole organism, compatible with immunohistochemistry and in situ hybridization is described.

Proteomics and pulse azidohomoalanine labeling of newly synthesized proteins: what are the potential applications?

The development of mass spectrometry (MS) based AHA strategies have the potential to measure proteins involved in immune response, secretome, gut microbiome, and proteostasis as well as their potential for clinical uses.

Identification and Quantitation of Newly Synthesized Proteins in Escherichia coli by Enrichment of Azidohomoalanine-labeled Peptides with Diagonal Chromatography

This work demonstrates for the first time the use of a bioorthogonal amino acid for proteome-wide detection of changes in the amounts of proteins synthesized during a brief period upon variations in cellular growth conditions.

Identification and Quantitation of Newly Synthesized Proteins in Escherichia coli by Enrichment of Azidohomoalanine-labeled Peptides with Diagonal Chromatography□S

This work demonstrates for the first time the use of a bioorthogonal amino acid for proteome-wide detection of changes in the amounts of proteins synthesized during a brief period upon variations in cellular growth conditions.

Enhancing Comprehensive Analysis of Newly Synthesized Proteins Based on Cleavable Bioorthogonal Tagging.

A novel method for enriching newly synthesized peptides by click chemistry followed by release of enriched peptides via tryptic digestion based on cleavable bioorthogonal tagging (CBOT) that can quantify NSPs when coupling a pair of isotope-labeled azidohomoalanine with decent reproducibility is designed.

Bio-orthogonal labeling as a tool to visualize and identify newly synthesized proteins in Caenorhabditis elegans

This protocol circumvents this limitation by identifying de novo–synthesized proteins via the incorporation of the chemically modifiable azidohomoalanine instead of the natural amino acid methionine in the nascent protein, followed by facilitating the visualization of the resulting labeled proteins in situ.

Comparative analysis of cleavable azobenzene-based affinity tags for bioorthogonal chemical proteomics.

Direct Detection of Biotinylated Proteins by Mass Spectrometry

DiDBiT improves the direct detection of biotin-tagged newly synthesized peptides more than 20-fold compared to conventional methods and demonstrates the MS detection of newly synthesize proteins labeled in vivo in the rodent nervous system with unprecedented temporal resolution as short as 3 h.

Identification and quantification of newly synthesized proteins translationally regulated by YB-1 using a novel Click-SILAC approach.

In situ visualization of newly synthesized proteins in environmental microbes using amino acid tagging and click chemistry

A direct comparison of anabolic activity using BONCAT and stable isotope labelling by nano-scale secondary ion mass spectrometry for individual cells within a sediment-sourced enrichment culture showed concordance between AHA-positive cells and 15N enrichment.
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