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… 

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.

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.

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.
...

References

SHOWING 1-10 OF 46 REFERENCES

Selective identification of newly synthesized proteins in mammalian cells using bioorthogonal noncanonical amino acid tagging (BONCAT).

This paper reports the selective purification and identification of 195 metabolically labeled proteins with multidimensional liquid chromatography in-line with tandem MS and says the identified proteins, synthesized in a 2-h window, possess a broad range of biochemical properties and span most functional gene ontology categories.

A tagging-via-substrate technology for detection and proteomics of farnesylated proteins.

TAS technology can be extended to other posttranslational modifications, such as geranylgeranylation and myristoylation, thus providing powerful tools for detection, quantification, and proteomic analysis of posttranslationally modified proteins.

Stable Isotope Labeling by Amino Acids in Cell Culture, SILAC, as a Simple and Accurate Approach to Expression Proteomics*

SILAC is a simple, inexpensive, and accurate procedure that can be used as a quantitative proteomic approach in any cell culture system and is applied to the relative quantitation of changes in protein expression during the process of muscle cell differentiation.

Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry

Comparison of the HMS-PCI data set with interactions reported in the literature revealed an average threefold higher success rate in detection of known complexes compared with large-scale two-hybrid studies.

Multiplexed Protein Quantitation in Saccharomyces cerevisiae Using Amine-reactive Isobaric Tagging Reagents*S

It is found that inactivation of Upf1p and Xrn1p causes common as well as unique effects on protein expression, and the use of 4-fold multiplexing to enable relative protein measurements simultaneously with determination of absolute levels of a target protein using synthetic isobaric peptide standards.

Status of complete proteome analysis by mass spectrometry: SILAC labeled yeast as a model system

Advanced mass spectrometry methods can unambiguously identify more than 2,000 proteins in a single proteome, including very low abundant ones, andSubstantially increased coverage of the yeast proteome appears feasible with further development in software and instrumentation.

Quantitative analysis of complex protein mixtures using isotope-coded affinity tags

An approach for the accurate quantification and concurrent sequence identification of the individual proteins within complex mixtures based on isotope-coded affinity tags and tandem mass spectrometry is described.

Analysis of quantitative proteomic data generated via multidimensional protein identification technology.

The method described provides an accurate way to undertake a large-scale quantitative proteomic study via multidimensional protein identification technology (MudPIT) and demonstrates the dynamic range of the system by analyzing a 1:1, 5:1 and 10:1 data set using the soluble portion from S. cerevisiae grown in either 14N or 15N-enriched minimal media.

Incorporation of azides into recombinant proteins for chemoselective modification by the Staudinger ligation

It is shown that proteins containing azidohomoalanine can be selectively modified in the presence of other cellular proteins by means of Staudinger ligation with triarylphosphine reagents and incorporation of azide-functionalized amino acids into proteins in vivo provides opportunities for protein modification under native conditions and selective labeling of proteins in the intracellular environment.