Site-Selective Synthetic Acylation of a Target Protein in Living Cells Promoted by a Chemical Catalyst/Donor System

Hamajima, Wataru; Fujimura, Akiko; Fujiwara, Yusuke; Yamatsugu, Kenzo; Kawashima, Shinichi; Kanai, Motomu

doi:10.1021/acschembio.9b00102

Cited by 18 publications

(16 citation statements)

References 29 publications

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“…We used two types of LANA-DSH catalysts, LANA 5-15 -DSH 1 and LANA 5-22 -DSH 2, and NAC-Ac 3 as a cell-permeable acetyl donor (Figure 1b). 12 As reported previously, 10 these catalysts efficiently promoted acetylation of H2BK120 in recombinant nucleosomes ( Figure S1). However, we could not detect H2BK120 acetylation by the catalyst system in living cells.…”

supporting

confidence: 73%

“…We then synthesized chemical catalysts conjugated with PEG-LANA ligands (Figure 3a). As the catalyst moiety, we used DSSMe, a pro-drugged DSH catalyst, 12 which is reduced to the active DSH form by biocompatible reductants, such as glutathione (GSH) or tris(2-carboxyethyl)phosphine (TCEP), prior to the reaction. The acetylation activity of the three catalysts (PEG 550 -LANA-DSSMe 11, PEG 750 -LANA-DSSMe 12, and PEG 2000 -LANA-DSSMe 13) was first examined in test tubes containing recombinant nucleosomes as substrates.…”

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confidence: 99%

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Live-Cell Epigenome Manipulation by Synthetic Histone Acylation Catalyst System

Fujiwara¹,

Yamanashi²,

Sato³

et al. 2020

Preprint

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<div><div><div><p>Chemical modifications of histones play a pivotal role in the epigenome and regulation of gene expression, and their abnormality is tightly linked to numerous disease states in humans. Therefore, chemical tools to manipulate epigenome hold promise for both therapy and the elucidation of epigenetic mechanisms. We previously developed the chemical catalyst LANA-DSH, which binds to nucleosomes via a LANA peptide ligand, and selectively acylates proximal histone H2BK120 to the catalyst moiety by acti- vating acyl-CoAs. Thus far, however, histone acylation by a chem- ical catalyst system in living cells has not yet been demonstrated. Here, we report a chemical catalyst system, composed of a nucleo- some-binding catalyst (PEG-LANA-DSH) and a cell-permeable thioester acyl donor (NAC-acyl), that can promote regioselective lysine acylation of histones in living cells. Whereas LANA-DSH is rapidly decomposed in cells, attachment of polyethylene glycol (PEG) to the LANA moiety can prevent this undesired degradation. Increasing the size of PEG conferred LANA with greater in-cell stability, but reduced catalytic activity, indicating that there is an optimum PEG length balancing stability and catalytic activity. The optimized PEG-LANA-DSH catalyst 11 efficiently promoted H2BK120 acetylation in living cells, which subsequently sup- pressed ubiquitination of H2BK120, a mark regulating various chromatin functions, such as transcription and DNA damage re- sponse. Thus, our chemical catalyst system will be useful as a unique tool to manipulate the epigenome for therapeutic purposes or further understanding epigenetic mechanisms.</p></div></div></div>

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confidence: 73%

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confidence: 99%

Live-Cell Epigenome Manipulation by Synthetic Histone Acylation Catalyst System

Fujiwara¹,

Yamanashi²,

Sato³

et al. 2020

Preprint

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“…In another principally similar report, biotin‐tethered diazotransfer reagent binds selectively to a protein for modification of a proximal lysine residue . Kanai and co‐workers demonstrated the efficacy of trimethoprim‐tethered nucleophilic catalyst for selective targeting of eDHFR in living cells . Also, Li and co‐workers reported sulfonium tethered peptide‐ligand directed modification of a Cys .…”

Section: Single‐site Labeling Of Native Proteinsmentioning

confidence: 94%

“…[121] Kanai and coworkers demonstrated the efficacy of trimethoprim-tethered nucleophilic catalyst for selective targeting of eDHFR in living cells. [122] Also, Li and co-workers reported sulfonium tethered peptide-ligand directed modification of a Cys. [123] Ball and coworkers demonstrated that a metallopeptide could provide proximity-driven catalysis for site-selective labeling of a protein.…”

Section: Ligand Directed Modificationsmentioning

confidence: 99%

Chemical Methods for Selective Labeling of Proteins

et al. 2019

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Protein bioconjugation poses outstanding questions of selectivity to the organic transformations. Besides, the presence of a pool of functional groups in the structural outfit of a protein brings its own set of characteristics. In this minireview, we highlight the challenges faced by a chemical transformation to deliver selectivity in the modification of proteins. The examples of pre‐engineered proteins outline the attributes associated with chemoselectivity and chemical orthogonality. Building on this foundation, we discuss the complexity of site‐selectivity in the single‐site modification of native proteins. The gradual evolution of chemical methods while addressing the challenges associated with different amino acids are outlined. The modular methods for labeling a residue independent of its reactivity order closes the discussion.

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“…1A). In contrast to the oligoDMAP-based catalysts, ligand-conjugated DSH catalysts can acetylate target proteins in living cells when the ligands are stable and functional in the cells (14). Biologically significant protein acetylation in living cells, such as histone acetylation, however, was not achieved yet.…”

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confidence: 99%

Live-cell epigenome manipulation by synthetic histone acetylation catalyst system

Fujiwara

Yamanashi

Fujimura

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

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Chemical modifications of histones, such as lysine acetylation and ubiquitination, play pivotal roles in epigenetic regulation of gene expression. Methods to alter the epigenome thus hold promise as tools for elucidating epigenetic mechanisms and as therapeutics. However, an entirely chemical method to introduce histone modifications in living cells without genetic manipulation is unprecedented. Here, we developed a chemical catalyst, PEG-LANA-DSSMe 11, that binds with nucleosome’s acidic patch and promotes regioselective, synthetic histone acetylation at H2BK120 in living cells. The size of polyethylene glycol in the catalyst was a critical determinant for its in-cell metabolic stability, binding affinity to histones, and high activity. The synthetic acetylation promoted by 11 without genetic manipulation competed with and suppressed physiological H2B ubiquitination, a mark regulating chromatin functions, such as transcription and DNA damage response. Thus, the chemical catalyst will be a useful tool to manipulate epigenome for unraveling epigenetic mechanisms in living cells.

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Site-Selective Synthetic Acylation of a Target Protein in Living Cells Promoted by a Chemical Catalyst/Donor System

Cited by 18 publications

References 29 publications

Live-Cell Epigenome Manipulation by Synthetic Histone Acylation Catalyst System

Live-Cell Epigenome Manipulation by Synthetic Histone Acylation Catalyst System

Chemical Methods for Selective Labeling of Proteins

Live-cell epigenome manipulation by synthetic histone acetylation catalyst system

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