Isotope-coded affinity tags (ICATs) are valuable tools for mass spectrometry-based quantitative proteomics, in particular, for comparison of protein (cysteine-residue) thiol oxidation state in normal, stressed, and diseased tissue. However, the iodoacetamido electrophile used in most commercial ICATs suffers from poor thiol-selectivity and modest rates of adduct formation, which can lead to spurious results. Hence, we designed and synthesized three ICATs containing thiol-selective -alkylmaleimide electrophiles (isotope-coded maleimide affinity tags = ICMATs) and assessed these as mass spectrometry probes for ratiometric analysis of lysozyme and muscle proteomes. Two ICMAT pairs containing butylene/D-butylene linkers were effective MS probes, but not ideal for typical proteomics workflows, because peptides bearing these tags frequently did not coelute with HPLC. A switch to a phenylene/C-phenylene linker solved this issue without compromising the efficiency of adduct formation.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1021/acs.bioconjchem.1c00206 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
The spurdog (Squalus acanthias Linnaeus, 1758) is a globally distributed squaliform shark that has historically been overfished but is now recovering in the northeast Atlantic. Data series on spurdog movement and habitat use have been somewhat limited to research surveys due to challenges associated with electronic tagging. Here, we offer a revised attachment method for externally attached pop-up satellite archival tags that was successful in long-term deployments on pregnant females.
View Article and Find Full Text PDFLysine malonylation is a post-translational modification where a malonyl group, characterized by a negatively charged carboxylate, is covalently attached to the ε-amino side chain of lysine, influencing protein structure and function. Our laboratory identified Mak upregulation in cartilage under aging and obesity, contributing to osteoarthritis (OA). Current antibody-based detection methods face limitations in identifying Mak targets.
View Article and Find Full Text PDFDesign of a yeast SUMO tag to eliminate internal translation initiation.
Protein Sci
January 2025
Department of Chemistry and Biochemistry, The Ohio State University, Columbus, Ohio, USA.
After overexpression in a suitable host, recombinant protein purification often relies on affinity (e.g., poly-histidine) and solubility-enhancing (e.
View Article and Find Full Text PDFGenetically encoded intrabody probes for labeling and manipulating AMPA-type glutamate receptors.
Nat Commun
November 2024
Department of Pharmacology, University of Colorado School of Medicine, Anschutz Medical Campus, Aurora, CO, 80045, USA.
Tools for visualizing and manipulating protein dynamics in living cells are critical for understanding cellular function. Here we leverage recently available monoclonal antibody sequences to generate a set of affinity tags for labeling and manipulating AMPA-type glutamate receptors (AMPARs), which mediate nearly all excitatory neurotransmission in the central nervous system. These antibodies can be produced from heterologous cells for exogenous labeling applications or directly expressed in living neurons as intrabodies, where they bind their epitopes in the endoplasmic reticulum and co-traffic to the cell surface for visualization with cell impermeant fluorescent dyes.
View Article and Find Full Text PDFA colorimetric biosensor composed of split aptamers and mannan oligosaccharide nanozyme to monitor synthetic His-tagged food biomolecules.
Food Chem
February 2025
State Key Laboratory of Marine Food Processing and Safety Control, College of Food Science and Engineering, Ocean University of China, Qingdao 266404, PR China; Laboratory for Marine Drugs and Bioproducts, Qingdao Marine Science and Technology Center, Qingdao 266237, PR China; Qingdao Key Laboratory of Food Biotechnology, Qingdao 266404, PR China; Key Laboratory of Biological Processing of Aquatic Products, China National Light Industry, Qingdao, 266404, PR China.
Food synthetic biology is garnering increasing attention for its potential to generate bioactive components. His-tag is one of the most popular tags used in food synthetic biology. Herein, His-tag, His-tagged proteins, and His-tagged peptides were adopted as the model targets, and a commonly used biosensor was developed to monitor His-tagged food biomolecules, using split aptamers as specific recognition probes and nanozyme as the transduction element.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!