Selective identification of newly synthesized proteins in mammalian cells using bioorthogonal noncanonical amino acid tagging (BONCAT).
AuthorsSchuman EM
JournalProceedings of the National Academy of Sciences of the United States of America
PubMed ID16769897
In both normal and pathological states, cells respond rapidly to environmental cues by synthesizing new proteins. The selective identification of a newly synthesized proteome has been hindered by the basic fact that all proteins, new and old, share the same pool of amino acids and thus are chemically indistinguishable. We ... More
In situ visualization and dynamics of newly synthesized proteins in rat hippocampal neurons.
AuthorsSchuman EM
JournalNature neuroscience
PubMed ID20543841
Protein translation has been implicated in different forms of synaptic plasticity, but direct in situ visualization of new proteins is limited to one or two proteins at a time. Here we describe a metabolic labeling approach based on incorporation of noncanonical amino acids into proteins followed by chemoselective fluorescence tagging ... More
Two-color labeling of temporally defined protein populations in mammalian cells.
AuthorsTirrell DA
JournalBioorganic & medicinal chemistry letters
PubMed ID18774715
The proteome undergoes complex changes in response to disease, drug treatment, and normal cellular signaling processes. Characterization of such changes requires methods for time-resolved protein identification and imaging. Here, we describe the application of two reactive methionine (Met) analogues, azidohomoalanine (Aha) and homopropargylglycine (Hpg), to label two protein populations in ... More
Fluorescence visualization of newly synthesized proteins in mammalian cells.
AuthorsTirrell DA
JournalAngewandte Chemie (International ed. in English)
PubMed ID17036290
Noncanonical amino acid tagging enables the selective fluorescent visualization of newly synthesized proteins in mammalian cells (see the picture). Susceptibility to tagging is determined by the spatial and temporal character of the protein synthesis, thus providing a complement to methods which identify relevant members of the proteome. ... More
Spatial coupling of mTOR and autophagy augments secretory phenotypes.
AuthorsNarita M., et al
JournalScience (New York, N.Y.)
PubMed ID21512002
Protein synthesis and autophagic degradation are regulated in an opposite manner by mammalian target of rapamycin (mTOR), whereas under certain conditions it would be beneficial if they occurred in unison to handle rapid protein turnover. We observed a distinct cellular compartment at the trans side of the Golgi apparatus, the ... More
Two-color labeling of temporally defined protein populations in mammalian cells.
AuthorsBeatty KE, Tirrell DA,
JournalBioorg Med Chem Lett
PubMed ID18774715
'The proteome undergoes complex changes in response to disease, drug treatment, and normal cellular signaling processes. Characterization of such changes requires methods for time-resolved protein identification and imaging. Here, we describe the application of two reactive methionine (Met) analogues, azidohomoalanine (Aha) and homopropargylglycine (Hpg), to label two protein populations in ... More
Dynamic monitoring of newly synthesized proteomes: up-regulation of myristoylated protein kinase A during butyric acid induced apoptosis.
AuthorsLiu K, Yang PY, Na Z, Yao SQ,
JournalAngew Chem Int Ed Engl
PubMed ID21678537
Doubly charged: A double metabolic incorporation approach capable of proteome-wide profiling of post-translational modification dynamics on newly synthesized proteins has been developed (see scheme; blue box: methionine surrogate, orange diamond: PTM probe). This strategy reveals for the first time that up-regulation of myristoylated PKA protein is necessary for the occurrence ... More
Expanding the genetic code of Saccharomyces cerevisiae with methionine analogues.
AuthorsWiltschi B, Wenger W, Nehring S, Budisa N,
JournalYeast
PubMed ID19061186
We replaced the single N-terminal methionine in heterologously expressed human Cu/Zn superoxide dismutase with the non-canonical methionine analogues homopropargylglycine and norleucine in the yeast Saccharomyces cerevisiae. Our non-canonical amino acid incorporation protocol involves a two-step procedure. In the first step, the methionine auxotrophic yeast cells are accumulated in synthetic medium ... More
Selectivity and specificity of substrate binding in methionyl-tRNA synthetase.
AuthorsDatta D, Vaidehi N, Zhang D, Goddard WA,
JournalProtein Sci
PubMed ID15388861
The accuracy of in vivo incorporation of amino acids during protein biosynthesis is controlled to a significant extent by aminoacyl-tRNA synthetases (aaRS). This paper describes the application of the HierDock computational method to study the molecular basis of amino acid binding to the Escherichia coli methionyl tRNA synthetase (MetRS). Starting ... More
Processing of N-terminal unnatural amino acids in recombinant human interferon-beta in Escherichia coli.
AuthorsWang A, Winblade Nairn N, Johnson RS, Tirrell DA, Grabstein K,
JournalChembiochem
PubMed ID18098265
Incorporation of unnatural amino acids into recombinant proteins represents a powerful tool for protein engineering and protein therapeutic development. While the processing of the N-terminal methionine (Met) residues in proteins is well studied, the processing of unnatural amino acids used for replacing the N-terminal Met remains largely unknown. Here we ... More
Click chemistry and bioorthogonal reactions: unprecedented selectivity in the labeling of biological molecules.
AuthorsBest MD,
JournalBiochemistry
PubMed ID19485420
In recent years, a number of bioorthogonal reactions have been developed, exemplified by click chemistry, that enable the efficient formation of a specific product, even within a highly complex chemical environment. While the exquisite selectivity and reliability of these transformations have led to their broad application in diverse research areas, ... More
In situ visualization and dynamics of newly synthesized proteins in rat hippocampal neurons.
AuthorsDieterich DC, Hodas JJ, Gouzer G, Shadrin IY, Ngo JT, Triller A, Tirrell DA, Schuman EM,
JournalNat Neurosci
PubMed ID20543841
Protein translation has been implicated in different forms of synaptic plasticity, but direct in situ visualization of new proteins is limited to one or two proteins at a time. Here we describe a metabolic labeling approach based on incorporation of noncanonical amino acids into proteins followed by chemoselective fluorescence tagging ... More
Functionally significant metabolic differences between B and T lymphocyte lineages.
Authors
JournalImmunology
PubMed ID31318442
Biofunctionalized aligned microgels provide 3D cell guidance to mimic complex tissue matrices.
Authors
JournalBiomaterials
PubMed ID29459322
Rapid Screening and Identification of Living Pathogenic Organisms via Optimized Bioorthogonal Non-canonical Amino Acid Tagging.
Authors
JournalCell Chem Biol
PubMed ID28757183
Neighboring cells override 3D hydrogel matrix cues to drive human MSC quiescence.
Authors
JournalBiomaterials
PubMed ID29852376
Live-cell imaging of alkyne-tagged small biomolecules by stimulated Raman scattering.
Authors
JournalNat Methods
PubMed ID24584195
Hypusination Orchestrates the Antimicrobial Response of Macrophages.
Authors
JournalCell Rep
PubMed ID33326776
Bi-directional cell-pericellular matrix interactions direct stem cell fate.
Authors
JournalNat Commun
PubMed ID30282987
Adaptive Protein Translation by the Integrated Stress Response Maintains the Proliferative and Migratory Capacity of Lung Adenocarcinoma Cells.
Authors
JournalMol Cancer Res
PubMed ID31551255
A chemical screen for modulators of mRNA translation identifies a distinct mechanism of toxicity for sphingosine kinase inhibitors.