Biological Sciences Division
Published November 25, 2024 | Version v1
Journal article Open

Circular Engineered Sortase for Interrogating Histone H3 in Chromatin

Creators

  • 1. Brigham and Women's Hospital
  • 2. Washington University in St. Louis
  • 3. University of Pennsylvania
  • 4. University of Leicester
  • 5. New York University
  • 6. University of Chicago

Description

Reversible modification of the histone H3 N-terminal tail is critical in regulating the chromatin structure, gene expression, and cell states, while its dysregulation contributes to disease pathogenesis. Understanding the crosstalk between H3 tail modifications in nucleosomes constitutes a central challenge in epigenetics. Here, we describe an engineered sortase transpeptidase, cW11, that displays highly favorable properties for introducing scarless H3 tails onto nucleosomes. This approach significantly accelerates the production of both symmetrically and asymmetrically modified nucleosomes. We demonstrate the utility of asymmetrically modified nucleosomes produced in this way in dissecting the impact of multiple modifications on eraser enzyme processing and molecular recognition by a reader protein. Moreover, we show that cW11 sortase is very effective at cutting and tagging histone H3 tails from endogenous histones, facilitating multiplex "cut-and-paste" middle-down proteomics with tandem mass tags. This cut-and-paste proteomics approach permits the quantitative analysis of histone H3 modification crosstalk after treatment with different histone deacetylase inhibitors. We propose that these chemoenzymatic tail isolation and modification strategies made possible with cW11 sortase will broadly power epigenetic discovery and therapeutic development.

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whedon-et-al-2024-circular-engineered-sortase-for-interrogating-histone-h3-in-chromatin.pdf

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Additional details

Identifiers

DOI
10.1021/jacs.4c12585
Other
oai:uchicago.tind.io:14159

Funding

National Institutes of Health
GM149229
National Science Foundation
2127882
National Institutes of Health
P01CA196539
National Institutes of Health
R01HD106051
Wellcome Trust
Investigator Award
University of Chicago
Nancy and Leonard Florsheim
National Institutes of Health
AR078555
National Institutes of Health
CA251677
National Institutes of Health
2R01GM115882-06
National Institutes of Health
1R01CA266978-01
National Institutes of Health
1R01GM144547-01A1
American Heart Association
Postdoctoral Fellowship
Charles King Trust
Postdoctoral Fellowship
Thermo Scientific Tandem Mass Tag Systems
Research Award

UChicago Information

Division(s)
Biological Sciences Division
Department(s)
Ben May Department for Cancer Research