Giordano Proietti

University of Southern Denmark

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Jasmin Mecinović

University of Southern Denmark

Yali Wang

Guangzhou Medical University

Tjie Kok

University of Surabaya

Jordi C. J. Hintzen

University of Gothenburg

Abbas H. K. Al Temimi

Radboud University Nijmegen

Frank J. Dekker

University of Groningen

Paul B. White

Radboud University Nijmegen

Gerrit J. Poelarends

University of Groningen

Hong Guo

University of Tennessee at Knoxville

Ping Qian

Shenzhen Third People’s Hospital

Cooperative Institutions

Radboud University Nijmegen

University of Groningen

Radboud Institute for Molecular Life Sciences

Radboud University Medical Center

University of Southern Denmark

Odense Municipality

Shandong Agricultural University

Huazhong University of Science and Technology

University of Oxford

Pharmaceutical Biotechnology (Czechia)

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Fine-tuning of lysine side chain modulates the activity of histone lysine methyltransferases

Scientific Reports (2020)

Abbas H. K. Al Temimi Jona Merx Christian J. van Noortwijk Giordano Proietti Romano Buijs

Histone lysine methyltransferases (KMTs) play an important role in epigenetic gene regulation and have emerged as promising targets for drug discovery. However, the scope and limitation of KMT catalysis on substrates possessing substituted lysine side chains remain insufficiently explored. Here, we identify new unnatural lysine analogues as substrates for human methyltransferases SETD7, SETD8, G9a and GLP. Two synthetic amino acids that possess a subtle modification on the lysine side chain, namely oxygen at the γ position (KO, oxalysine) and nitrogen at the γ position (KN, azalysine) were incorporated into histone peptides and tested as KMTs substrates. Our results demonstrate that these lysine analogues are mono-, di-, and trimethylated to a different extent by trimethyltransferases G9a and GLP. In contrast to monomethyltransferase SETD7, SETD8 exhibits high specificity for both lysine analogues. These findings are important to understand the substrate scope of KMTs and to develop new chemical probes for biomedical applications.

10.1038/s41598-020-78331-0

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Citations (6)

Structure-activity relationships for binding of 4-substituted triazole-phenols to macrophage migration inhibitory factor (MIF)

European Journal of Medicinal Chemistry (2019)

Zhangping Xiao Marieke Fokkens Deng Chen Tjie Kok Giordano Proietti

Macrophage migration inhibitory factor (MIF) is a versatile protein that plays a role in inflammation, autoimmune diseases and cancers. Development of novel inhibitors will enable further exploration of MIF as a drug target. In this study, we investigated structure-activity relationships of MIF inhibitors using a MIF tautomerase activity assay to measure binding. Importantly, we notified that transition metals such as copper (II) and zinc (II) interfere with the MIF tautomerase activity under the assay conditions applied. EDTA was added to the assay buffer to avoid interference of residual heavy metals with tautomerase activity measurements. Using these assay conditions the structure-activity relationships for MIF binding of a series of triazole-phenols was explored. The most potent inhibitors in this series provided activities in the low micromolar range. Enzyme kinetic analysis indicates competitive binding that proved reversible. Binding to the enzyme was confirmed using a microscale thermophoresis (MST) assay. Molecular modelling was used to rationalize the observed structure-activity relationships. The most potent inhibitor 2d inhibited proliferation of A549 cells in a clonogenic assay. In addition, 2d attenuated MIF induced ERK phosphorylation in A549 cells. Altogether, this study provides insights in the structure-activity relationships for MIF binding of triazole-phenols and further validates this class of compounds as MIF binding agents in cell-based studies.

Microscale Thermophoresis

Structure–activity relationship

10.1016/j.ejmech.2019.111849

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Citations (26)

Examining sterically demanding lysine analogs for histone lysine methyltransferase catalysis

Scientific Reports (2020)

Abbas H. K. Al Temimi Vu Tran Ruben S. Teeuwen Arthur J. Altunc Helene I. V. Amatdjais‐Groenen

Abstract Methylation of lysine residues in histone proteins is catalyzed by S -adenosylmethionine (SAM)-dependent histone lysine methyltransferases (KMTs), a genuinely important class of epigenetic enzymes of biomedical interest. Here we report synthetic, mass spectrometric, NMR spectroscopic and quantum mechanical/molecular mechanical (QM/MM) molecular dynamics studies on KMT-catalyzed methylation of histone peptides that contain lysine and its sterically demanding analogs. Our synergistic experimental and computational work demonstrates that human KMTs have a capacity to catalyze methylation of slightly bulkier lysine analogs, but lack the activity for analogs that possess larger aromatic side chains. Overall, this study provides an important chemical insight into molecular requirements that contribute to efficient KMT catalysis and expands the substrate scope of KMT-catalyzed methylation reactions.

Histone Methylation

10.1038/s41598-020-60337-3

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Citations (10)

CCDC 1575884: Experimental Crystal Structure Determination

The Cambridge Structural Database (2018)

Tjie Kok Hannah Wapenaar Kan Wang Constantinos G. Neochoritis Tryfon Zarganes‐Tzitzikas

An entry from the Cambridge Structural Database, the world’s repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.

10.5517/ccdc.csd.cc1pwtzc

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Investigation of in vitro histone H3 glycosylation using H3 tail peptides

Scientific Reports (2022)

Jona Merx Jordi C. J. Hintzen Giordano Proietti Hidde Elferink Yali Wang

Abstract Posttranslational modifications (PTMs) on histone tails regulate eukaryotic gene expression by impacting the chromatin structure and by modulating interactions with other cellular proteins. One such PTM has been identified as serine and threonine glycosylation, the introduction of the ß- N -acetylglucosamine (GlcNAc) moiety on histone H3 tail at position Ser10 and Thr32. The addition of the ß- O -GlcNAc moiety on serine or threonine residues is facilitated by the O -GlcNAc transferase (OGT), and can be removed by the action of O -GlcNAcase (OGA). Conflicting reports on histone tail GlcNAc modification in vivo prompted us to investigate whether synthetic histone H3 tail peptides in conjunction with other PTMs are substrates for OGT and OGA in vitro. Our enzymatic assays with recombinantly expressed human OGT revealed that the unmodified and PTM-modified histone H3 tails are not substrates for OGT at both sites, Ser10 and Thr32. In addition, full length histone H3 was not a substrate for OGT. Conversely, our work demonstrates that synthetic peptides containing the GlcNAc functionality at Ser10 are substrates for recombinantly expressed human OGA, yielding deglycosylated histone H3 peptides. We also show that the catalytic domains of human histone lysine methyltransferases G9a, GLP and SETD7 and histone lysine acetyltransferases PCAF and GCN5 do somewhat tolerate glycosylated H3Ser10 close to lysine residues that undergo methylation and acetylation reactions, respectively. Overall, this work indicates that GlcNAcylation of histone H3 tail peptide in the presence of OGT does not occur in vitro.

10.1038/s41598-022-21883-0

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Substrate Scope for Human Histone Lysine Acetyltransferase KAT8

International Journal of Molecular Sciences (2021)

Giordano Proietti Yali Wang Chiara Punzo Jasmin Mecinović

Biomedically important histone lysine acetyltransferase KAT8 catalyses the acetyl coenzyme A-dependent acetylation of lysine on histone and other proteins. Here, we explore the ability of human KAT8 to catalyse the acetylation of histone H4 peptides possessing lysine and its analogues at position 16 (H4K16). Our synthetic and enzymatic studies on chemically and structurally diverse lysine mimics demonstrate that KAT8 also has a capacity to acetylate selected lysine analogues that possess subtle changes on the side chain and main chain. Overall, this work highlights that KAT8 has a broader substrate scope beyond natural lysine, and contributes to the design of new chemical probes targeting KAT8 and other members of the histone lysine acetyltransferase (KAT) family.

Histone acetyltransferase

Acetyltransferases

Histone H4

10.3390/ijms22020846

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Citations (12)

Discovery of chromenes as inhibitors of macrophage migration inhibitory factor

Bioorganic & Medicinal Chemistry (2017)

Tjie Kok Hannah Wapenaar Kan Wang Constantinos G. Neochoritis Tryfon Zarganes‐Tzitzikas

CD74

10.1016/j.bmc.2017.12.032

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Citations (17)

γ-Thialysine versus Lysine: An Insight into the Epigenetic Methylation of Histones

Bioconjugate Chemistry (2019)

Abbas H. K. Al Temimi Renate van der Wekken-de Bruijne Giordano Proietti Hong Guo Ping Qian

Biomedicinally important histone lysine methyltransferases (KMTs) transfer a methyl group from S-adenosylmethionine to lysine residues in histones and other proteins. Here, we report comparative studies on epigenetic methylation of lysine and γ-thialysine, the simplest cysteine-derived lysine analog, which can be introduced to histone peptides and histone proteins via site-specific bioconjugation-based cysteine alkylation. Enzyme assays and computational studies demonstrate that human KMTs catalyze efficient methylation of histones that possess γ-thialysine. This work provides a molecular basis for the application of γ-thialysine for biomolecular studies of intact histones and the nucleosome assembly.

Histone Methylation

10.1021/acs.bioconjchem.9b00313

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Citations (16)

Exploring the Histone Acylome through Incorporation of γ-Thialysine on Histone Tails

Bioconjugate Chemistry (2020)

Giordano Proietti Giorgio Rainone Jordi C. J. Hintzen Jasmin Mecinović

Histone lysine acetyltransferases (KATs) catalyze the transfer of the acetyl group from acetyl Coenzyme A to lysine residues in histones and nonhistone proteins. Here, we report biomolecular studies on epigenetic acetylation and related acylation reactions of lysine and γ-thialysine, a cysteine-derived lysine mimic, which can be site-specifically introduced to histone peptides and histone proteins. Enzyme assays demonstrate that human KATs catalyze an efficient acetylation and propionylation of histone peptides that possess lysine and γ-thialysine. Enzyme kinetics analyses reveal that lysine- and γ-thialysine-containing histone peptides exhibit indistinguishable Km values, whereas small differences in kcat values were observed. This work highlights that γ-thialysine may act as a representative and easily accessible lysine mimic for chemical and biochemical examinations of post-translationally modified histones.

Histone H4

Histone octamer

10.1021/acs.bioconjchem.0c00012

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Citations (13)

Effect of lysine side chain length on histone lysine acetyltransferase catalysis

Scientific Reports (2020)

Giordano Proietti Yali Wang Giorgio Rainone Jasmin Mecinović

Abstract Histone lysine acetyltransferase (KAT)-catalyzed acetylation of lysine residues in histone tails plays a key role in regulating gene expression in eukaryotes. Here, we examined the role of lysine side chain length in the catalytic activity of human KATs by incorporating shorter and longer lysine analogs into synthetic histone H3 and H4 peptides. The enzymatic activity of MOF, PCAF and GCN5 acetyltransferases towards histone peptides bearing lysine analogs was evaluated using MALDI-TOF MS assays. Our results demonstrate that human KAT enzymes have an ability to catalyze an efficient acetylation of longer lysine analogs, whereas shorter lysine analogs are not substrates for KATs. Kinetics analyses showed that lysine is a superior KAT substrate to its analogs with altered chain length, implying that lysine has an optimal chain length for KAT-catalyzed acetylation reaction.

PCAF

Acetyltransferases

Histone acetyltransferase

Histone H4

10.1038/s41598-020-69510-0

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Citations (12)