Biol. Chem. 2021; ▪▪▪(▪▪▪): 1–44 Review Anna-Theresa Blasl, Sabrina Schulze, Chuan Qin, Leonie G. Graf, Robert Vogt and Michael Lammers* Post-translational lysine ac(et)ylation in health, ageing and disease https://doi.org/10.1515/hsz-2021-0139 protein function, how it is regulated enzymatically and non- Received February 8, 2021; accepted June 18, 2021; enzymatically, how a dysfunction in this post-translational published online August 23, 2021 machinery contributes to disease development. A focus is set on sirtuins and lysine acyltransferases as these are direct Abstract: The acetylation/acylation (ac(et)ylation) of lysine sensors and mediators of the cellular metabolic state. side chains is a dynamic post-translational modification Finally, this review highlights technological advances to (PTM) regulating fundamental cellular processes with im- study lysine ac(et)ylation. plications on the organisms’ ageing process: metabolism, transcription, translation, cell proliferation, regulation of Keywords: ageing; KDAC; longevity; lysine acetylation; the cytoskeleton and DNA damage repair. First identified to lysine acetyltransferases; lysine deacetylases; sirtuins; occur on histones, later studies revealed the presence of synthetic biology. lysine ac(et)ylation in organisms of all kingdoms of life, in proteins covering all essential cellular processes. A remarkable finding showed that the NAD+-dependent Introduction sirtuin deacetylase Sir2 has an impact on replicative lifespan in Saccharomyces cerevisiae suggesting that lysine acetyla- The human genome encodes approximately 20,000–25,000 tion has a direct role in the ageing process. Later studies proteins (International Human Genome Sequencing fi fi identi ed sirtuins as mediators for bene cial effects of 2004). The diversity of this proteome can be substantially ’ caloric/dietary restriction on the organisms health- or life- enlarged by processes such as alternative splicing and span. However, the molecular mechanisms underlying post-translational modifications (PTMs). PTMs range from these effects are only incompletely understood. Progress in attachment of chemical groups such as observed in mass-spectrometry, structural biology, synthetic and semi- phosphorylation of serine, threonine or tyrosine side synthetic biology deepened our understanding of this PTM. chains dynamically regulated by kinases and phospha- This review summarizes recent developments in the tases, to attachment of small proteins such as ubiquitin fi research eld. It shows how lysine ac(et)ylation regulates for regulation of protein turnover, subcellular localization and signal transduction. Moreover, proteolytic processing of proteins is a well-known PTM as exemplified by the Anna-Theresa Blasl and Sabrina Schulze are contributed equally to activation of coagulation factors or the activation of this article. digestive enzymes from inactive zymogens. Proteins can *Corresponding author: Michael Lammers, Department Synthetic and be lipidated to regulate membrane binding/recruitment Structural Biochemistry, Institute for Biochemistry, University of by forming irreversible thioethers between cysteine Greifswald, Felix-Hausdorff-Str. 4, D-17487 Greifswald, Germany, side chains and prenyl-groups or by forming reversible E-mail: [email protected]. https://orcid.org/ thioesters by attachment of fatty acids (palmitoylation, 0000-0003-4168-4640 Anna-Theresa Blasl, Sabrina Schulze, Chuan Qin, Leonie G. Graf myristoylation) to cysteine side chains (Chen et al. 2018). and Robert Vogt, Department Synthetic and Structural Biochemistry, Apart from reversible lipidation on cysteine side chains, Institute for Biochemistry, University of Greifswald, Felix-Hausdorff- lysine side chains can undergo reversible lipidation/ Str. 4, D-17487 Greifswald, Germany, E-mail: anna- acylation. [email protected] (A.-T. Blasl), sabrina.schulze@uni- Lysine side chains can be modified by various post- greifswald.de (S. Schulze), [email protected] (C. Qin), translational modifications. Lysines can be modified by [email protected] (L.G. Graf), robert.vogt@uni- greifswald.de (R. Vogt). https://orcid.org/0000-0002-0122-5688 ubiquitination (Ub) or by attachment of a ubiquitin-like (S. Schulze) (Ubl) protein such as SUMO1-5, ISG15 or Nedd8 by Open Access. © 2021 Anna-Theresa Blasl et al., published by De Gruyter. This work is licensed under the Creative Commons Attribution 4.0 International License. 2 A.-T. Blasl et al.: Lysine ac(et)ylation in health, ageing and disease formation of an isopeptide bond between the ε-amino and YEATS domains (Figure 1(A)). We will discuss how group of the lysine side chain and the α-carboxyl group of lysine acetylation is involved in disease development and the terminal diGly motif of Ub/Ubl (Celen and Sahin 2020; will put an emphasis on recent technological advances to Swatek and Komander 2016). The ubiquitin can be removed study lysine ac(et)ylation. by deubiquitinases (DUBs). Moreover, lysine side chains can be modified by mono-/di-/tri-methylation, which alters the side chain sterically and enables the binding of Enzymatic regulation of lysine proteins containing specific methyl-lysine reader domains. ac(et)ylation by classical KDACs, Tudor domains enable binding to di- and trimethylated lysines, while MBT domains bind to mono-/di-methylated sirtuins and KATs lysines and PHD domains bind to various methylation 2+ states (mono-/di-/tri-methyl-lysine) (Wang et al. 2020a,b). Classical Zn -dependent lysine PHD domains are zinc containing domains that often occur deacetylases as tandem PHD domains. Chromodomains, PWWP- and WD domains bind to methylated lysine residues. All of After the discovery of lysine acetylation on histones in these domains target various methylated lysines in his- 1963, enzymatic activity to remove acetyl-groups was tone tails and are involved in chromatin remodeling and - discovered in calf thymus in 1969 (Inoue and Fujimoto regulation of gene expression. Lysine side chains can be 1969; Phillips 1963). However, the isolation of the first modified by acetylation and various other acylations lysine deacetylase HDAC1 took until 1996 (Taunton et al. such as the charged succinylation, malonylation or gluta- 1996). Later studies showed the presence of two lysine rylation and aliphatic acylations such as formylation, deacetylase families, which are structurally and based on propionylation, butyrylation, myristoylation or palmitoy- their primary sequences unrelated: 1.) classical histone/ lation (Figure 1(A)–(C)). Recently, lysine 2-hydrox lysine decacetylases (HDACs/KDACs; Zn2+-dependent yisobutyrylation occurring on histones with high stoichi- catalytic mechanism) and sirtuin deacetylases (sirtuins; ometry inducing structural changes in the chromatin was SIRT: silent information regulator; NAD+-dependent shown to regulate glycolysis and being catalyzed by the catalytic mechanism) (Seto and Yoshida 2014; Yoshida KAT p300 (Figure 1(C)) (Dai et al. 2014; Huang et al. et al. 2017). As the deacetylases were first characterized as 2018a,b, 2020). In contrast to mono-/di-/tri-methylation, histone deacetylases, the name was abbreviated with the acetylation/acylation of lysine side chains results in HDAC. However, as recent data show that these enzymes neutralization of the positive charge occurring at the lysine have many substrates besides histones they are also called side chain at physiological pH. lysine deacetylases (KDACs) (Kumar et al. 2021; Peng and Lysine acetylation was identified together with Seto 2011; Sadoul et al. 2011). Today it is known that 11 methylation by Phillips in 1963 to occur on histones and in classical deacetylases are encoded in the human genome 1964 Allfrey reported that acetylation and methylation of (Figure 2(A)) classified into three classes: class I (with histones regulates RNA synthesis, suggesting that it has HDAC1/2/3/8 showing sequence similarity to yeast Rpd3), an impact on gene expression (Allfrey and Mirsky 1964; class II (subclass IIa with HDAC4/5/7/9 and subclass IIb Allfrey et al. 1964; Phillips 1963). Today it is known that not with HDAC6/10 showing sequence similarity to yeast Hda1) only histones are lysine-acetylated, but in fact the number and class IV (with HDAC11 showing sequence similarity to of lysine ac(et)ylation sites rivals that of phosphorylation class I and class II enzymes) (Figure 2(A)) (de Ruijter et al. sites and thousands of proteins are acetylated in all cellular 2003; Haberland et al. 2009). These enzymes of classes I, II compartments and it is present in all kingdoms of life and IV are structurally similar and use a catalytic Zn2+ ion (Choudhary and Mann 2020; Choudhary et al. 2009, 2014; to hydrolyze the amide bond to remove acetyl-groups from Hansen et al. 2019; Kouzarides 2000; Lundby et al. 2012; the ε-amino group of lysine side chains (Figure 2(B) and Weinert et al. 2011, 2015, 2017, 2018). (C)) (Lombardi et al. 2011). This review highlights the physiological role of lysine The catalytic mechanism involves the coordination acetylation. We discuss the regulation of lysine acetylation and polarization of a catalytic water molecule by the mechanistically with respect to cellular metabolic path- catalytic Zn2+-ion (Figure 2(C)). This Zn2+-ion together with ways. It summarizes how lysine acetylation is regulated in a Tyr residue (HDAC8: Y306), which is replaced by a his- cells by writers, the lysine acetyltransferases (KATs), tidine in class IIa KDACs, also interact with the carbonyl erasers, the lysine