Fischle/Schwarzer

Establishing a Chemical Tool Box for Modified Nucleosomes and its Application for Analysis of the Histone Code

Nucleosomes constitute the basic structural unit of chromatin composed of histone proteins that form a scaffold with about 150 base pairs of DNA wrapped around. Chromatin compacts eukaryotic genomes and represents an important means for regulating gene activity which is mediated by a multitude of posttranslational modifications on the N-terminal tail regions of the histones. The exact nature of the complex cross-talk between histone modifications and gene activity remains unknown in part due to the lack of techniques for generating nucleosomes with defined modification patterns in large numbers.

Here we propose establishing a tool box for modified nucleosomes with defined modification patterns on histones H3 and H4. To this end we plan to generate ligation-ready nuclosomes with truncated histones H3 and H4, which can be orthogonally ligated with the corresponding synthetic tails by chemoselective ligation techniques.

To realize this plan we will establish an improved sortase-mediated ligation for histone H3 and protein trans-splicing based on the Ssp DnaB intein for H4 in the context of immobilized nucleosomes. A nucleosomal library with a complexity of 512 modification patterns should be synthesized and used for probing a set of modification-dependent histone binding proteins in a combinatorial type of fashion. Selected modified nucleosomes should be further used for interaction studies on a proteomic level. In addition we plan to use ligation-ready nucleosomes for installing various fluorophores at different histone sites and screen for ideal positioning of FRET donor and acceptor pairs for quantitative binding studies.

Dr. Wolfgang Fischle
Georg-August-Universität Göttingen

Tel.: +49 551 201 1340
Fax: +49 551 201 1197

Email Dr. Fischle

Prof. Dr. Dirk Schwarzer
Eberhard Karls Universität Tübingen

Tel.: +49 7071 29 - 73344
Fax: +49 7071 29 - 4815

Email Prof. Schwarzer

Publications within the SPP 1623 project

W. Fischle, D. Schwarzer
ACS Chemical Biology 2016, 11(3), 689-705 (Review)
Probing Chromatin-modifying enzymes with Chemical Tools.
Link to the article

A. Stützer, S. Liokatis, A. Kiesel, D. Schwarzer, R. Sprangers, J. Söding, P. Selenko, W. Fischle
Molecular Cell 2016, 61, 247-259
Modulations of DNA Contacts by Linker Histones and Post-translational Modifications Determine the Mobility and Modifiability of Nucleosomal H3 Tails.
Link to the article

A. Dose, J. Sindlinger, J. Bierlmeier, A. Bakirbas, K. Schulze-Osthoff, S. Einsele-Scholz, M. Hartl, F. Essmann, I. Finkemeier, D. Schwarzer
Angew. Chem. Int. Ed. 2016, 55, 1192-1195
Interrogating Substrate Selectivity and Composition of Endogenous Histone Deacetylase Complexes with Chemical Probes.
Link to the article

J. Seidel, C. Klockenbusch, D. Schwarzer
Chembiochem 2015, Ahead of print
Investigating deformylase and deacylase activity of mammalian and bacterial sirtuins.

W. Fischle, H. Mootz, D. Schwarzer
Curr. Opin. Chem. Biol. 2015, 28, 131-140 (Review)
Synthetic histone code.
Link to the article

O. Jost, A. Hanswillemenke, D. Schwarzer
Mol Biosyst. 2015, 11, 1820-1823
A miniaturized readout strategy for endogenous histone ceacetylase activity.
Link to the article

L. Schmohl, F.R. Wagner, M. Schümann, E. Krause, D. Schwarzer
Bioorg. Med. Chem. 2015, 15, 2883-2889
Semisynthesis and initial characterization of sortase A mutants containing selenocysteine and homocysteine.
Link to the article

L. Schmohl, D. Schwarzer
Curr. Opin. Chem. Biol. 2014, 22, 122-128
Sortase-mediated ligations for the site-specific modification of proteins
Link to the article

K.A. Gelato, M. Tauber, M.S. Ong, S. Winter. K. Hiragami-Hamada, J. Sindlinger, A. Lemak, Y. Bultsma, S. Houliston, D. Schwarzer, N. Divecha, C.H. Arrowsmith, W. Fischle
Mol. Cell 2014, 54, 905-919
Accessibility of Different Histone H3-Binding Domains of UHRF1 Is Allosterically Regulated by Phosphatidylinositol 5-Phosphate.
Link to the article

L. Schmohl, D. Schwarzer
J. Pept. Sci. 2014, 20, 145-151
Chemo-enzymatic three-fragment assembly of semisynthetic proteins.
Link to the article