Engineered bromodomains to explore the acetylproteome
Name
Yaffe_Engineered bromodomains.pdf
Size
701.42 KB
Format
Adobe PDF
Checksum (MD5)
f267878074933fecf34a90bd21b1527a
Author(s)
Del Rosario, Amanda M
Yaffe, Michael B
Gootenberg, Jonathan S
Bryson, Bryan D.
White, Forest M.
Date Issued
April 2015
Journal
PROTEOMICS
Publisher
Wiley Blackwell
Citation
Bryson, Bryan D. et al. “Engineered Bromodomains to Explore the Acetylproteome.” PROTEOMICS 15.9 (2015): 1470–1475.
Version
Author's final manuscript
Abstract
Mass spectrometry-based analysis of the acetylproteome has highlighted a role for acetylation in a wide array of biological processes including gene regulation, metabolism, and cellular signaling. To date, anti-acetyllysine antibodies have been used as the predominant affinity reagent for
enrichment of acetyllysine-containing peptides and proteins; however, these reagents suffer from high non-specific binding and lot-to-lot variability. Bromodomains represent potential affinity reagents for acetylated proteins and peptides, given their natural role in recognition of acetylated
sequence motifs in vivo. To evaluate their efficacy, we generated recombinant proteins representing all known yeast bromodomains. Bromodomain specificity for acetylated peptides was determined using degenerate peptide arrays, leading to the observation that different
bromodomains display a wide array of binding specificities. Despite their relatively weak affinity, we demonstrate the ability of selected bromodomains to enrich acetylated peptides from a complex biological mixture prior to mass spectrometric analysis. Finally, we demonstrate a method for improving the utility of bromodomain enrichment for mass spectrometry through engineering novel affinity reagents using combinatorial tandem bromodomain pairs.
MIT Department
Massachusetts Institute of Technology. Department of Biological Engineering
Massachusetts Institute of Technology. Department of Biology
Koch Institute for Integrative Cancer Research at MIT
Terms of Use
Creative Commons Attribution-Noncommercial-Share Alike
Persistent DSpace Link
DOI of Published Version
https://doi.org/10.1002/pmic.201400401
