Post-translational modifications alter the properties and activities of a protein through:

• Addition of chemical groups: Attaching molecules such as acetyl, phosphate, glycosyl, or methyl groups to specific amino acids.
• Complete protein degradation: Controlling protein lifespan and recycling cellular components.
• Proteolytic cleavage: Breaking down protein segments to activate or deactivate functions.

PTMs regulate essential cellular functions, enable signal transduction, and enhance proteome complexity by diversifying protein structures and roles. Advances in proteomics and bioinformatics have uncovered PTM cross-talk, highlighting its role in higher-order regulation and functional modulation of cellular processes.

Types of post-translational modifications

There are over 650 known types of PTMs, with some of the most common including phosphorylation, glycosylation, acetylation, methylation, and ubiquitination, which account for over 90% of reported modifications. Amino acids like lysine, cysteine, and serine undergo multiple PTMs, with phosphorylation on serine being the most frequently observed¹.

Enzymatic and non-enzymatic post-translational modifications

PTMs can be broadly either enzymatic (enzyme-catalyzed) or non-enzymatic. Enzymatic PTMs are modifications that require enzyme-mediated catalysis to modify specific amino acid residues on proteins. Enzymatic PTMs are tightly regulated by specific enzymes and signaling pathways.

• Methylation: Catalyzed by methyltransferases for the addition of methyl groups.
• Phosphorylation: Phosphate groups are added, and the reaction is catalyzed by kinases.
• Acetylation: Acetyltransferases-mediated addition of acetyl groups.
• Glycosylation: Glycosyltransferases-mediated addition of sugar molecules.
• SUMOylation: The addition of a small ubiquitin-like modifier (SUMO) under the action of three enzymes, E1, E2, and E3, respectively, that carry out activation, conjugation, and ligation.
• Ubiquitination involves three enzymes: ubiquitin-activating, E1; ubiquitin-conjugating, E2; and ubiquitin ligase, E3, to add ubiquitin to target proteins.
• Acetylation: Acetyl groups are added through acetylation under the action of acetyltransferases and deacetylases.
• Lipidation: Involves the action of palmitoyl transferases, N-myristoyltransferases, and depalmitoylation enzymes.

Non-enzymatic PTMs occur spontaneously through chemical reactions between amino acid side chains on proteins and reactive metabolites. These PTMs are often influenced by cellular stress, metabolic imbalances, and aging, leading to uncontrolled modifications. For example:

• Oxidation: Modification of amino acids, mediated by reactive oxygen species, leading to protein damage or dysfunction.
• Glycation: Reaction between sugars with amino acids, forming advanced glycation end-products (AGEs).
• Carbonylation: Amino acids are modified and involve reactive carbonyl compounds.
• Pyrophosphorylation: Addition of a phosphate group to a pre-existing phosphoserine residue. This non-enzymatic PTM is mediated by inositol pyrophosphates.

Covalent modifications

Post-translational modifications involve covalent bond changes in proteins, enhancing their diversity and playing a pivotal role in regulating enzyme activity, protein interactions, and cellular signaling.

Phosphorylation: Mechanism and impact on cell signaling

Protein phosphorylation is a crucial post-translational modification that functions as a molecular switch. In this process, kinases transfer a phosphate (PO₄³⁻) group from ATP to specific amino acids - such as serine, threonine, tyrosine, or histidine - while phosphatases reverse this modification by removing the phosphate group. This reversible process allows for the rapid regulation of protein function, influencing enzyme activity, membrane channels, and protein interactions in both prokaryotic and eukaryotic cells, in the nucleus but more commonly in the cytosol.

The most common type of phosphorylation is on serine/threonine residues, which is mediated by kinases such as mitogen-activated protein kinases (MAPKs), protein kinase A (PKA), and protein kinase C (PKC). Phosphorylation on serine/threonine residues is commonly involved in the regulation of key biological functions such as apoptosis, metabolism, stress responses, and cell cycle progression².

Although more common in bacterial two-component systems, histidine phosphorylation also controls metabolic enzymes and intracellular signaling in eukaryotes. Its volatility under physiological settings makes identification difficult, but its importance in eukaryotic signaling deserves more research.

Tyrosine phosphorylation is performed by non-receptor tyrosine kinases (NRTKs) and RTKs, and is critical for cell signaling, particularly in growth factors and immunological pathways (Figure 1).

This image is from an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Targeting RTK Signaling Pathways in Cancer. Cancers September 2015. 7(3):1758-1784. DOI: 10.3390/cancers7030860

Figure 1: Upon ligand binding, RTKs like vascular endothelial growth factor receptor (VEGFR) and EGFR activate downstream pathways, such as phosphatidylinositol 3-kinase-protein kinase B, Janus kinase-signal transducer and activator of transcription, and RAS-RAF-mitogen-activated protein kinase-extracellular-signal-regulated kinase, which regulate survival, differentiation, and proliferation of cells.

Threonine phosphorylation is the process of adding a phosphate group to the hydroxyl group of the threonine side chain. Numerous biological processes, including cell survival, cell cycle regulation, metabolism, and development, are impacted by threonine phosphorylation.

In glutamic acid phosphorylation, a phosphate group attaches to a glutamic acid residue, altering the charge distribution and potentially affecting cellular signaling, regulatory mechanisms, and protein interactions.

Phosphorylation of aspartic acid involves the addition of a phosphate group to an aspartic acid residue. Consequently, the charge distribution of the protein may be altered, potentially altering its structure and its function. Bacterial two-component signal transduction systems frequently exhibit this phosphorylation, which is an essential regulatory mechanism in response to environmental changes³.

Aberrant phosphorylation can lead to protein misfolding and aggregation, resulting in dysregulation of cellular functions and, ultimately, disease development. In Alzheimer’s disease, for example, hyperphosphorylated tau forms neurofibrillary tangles. The course of disease is aided by disrupted cell cycle regulation, apoptosis, and protein degradation. In addition, cancer, metabolic dysfunction, and compromised stress responses brought on by dysregulated kinase activity⁴.

Glycosylation: Types and importance in protein stability and immune response

Glycosylation, a complex and reversible post-translational modification, involves attaching sugar molecules to specific protein residues, primarily serine, threonine, asparagine, and tryptophan. It plays an important role in processes like:

• Protein folding
• Cell signaling
• Molecular trafficking

It can also affect the half-life of proteins in circulation.

Mechanisms and variants of glycosylation

N-linked glycosylation binds glycans to asparagine residues within a certain consensus sequence. The process begins in the endoplasmic reticulum (ER) with a lipid-linked oligosaccharide, which is subsequently carried to the protein and processed in the ER and Golgi apparatus.

O-linked glycosylation adds sugars to serine or threonine residues. In contrast to N-linked glycosylation, it mostly occurs in the Golgi apparatus and lacks a consensus sequence. The starting sugar, which is often N-acetylgalactosamine (GalNAc), is converted into complex glycans.

Other types of glycosylation include C-mannosylation, which adds mannose to tryptophan, and glypiation, which attaches glycosylphosphatidylinositol anchors to proteins, securing them to cell membranes⁵.

Glycosylation-dependent pathways

Glycosylation is necessary for numerous biological activities. Antibody glycosylation influences immunological stability and interactions. Glycan-binding proteins such as selectins facilitate cell-cell interaction and adhesion, directing leukocyte transport during inflammatory responses. Glycosylation also governs protein folding and quality control in the ER, where N-linked glycans signal chaperone proteins to fold properly and degrade misfolded proteins.

Dysregulated glycosylation

Aberrant glycosylation contributes to illness. In cancer, abnormal glycosylation accelerates tumor growth by encouraging invasion, immunological evasion, and cell detachment. Genetic glycan biosynthesis deficiencies result in congenital glycosylation disorders, which induce developmental and organ failure.

Glycosylation changes occur throughout neurodegeneration and aging, influencing disease progression. Protein aggregation is responsible for Alzheimer’s disease-related poor glycosylation of the amyloid precursor protein. Changes in immunoglobulin G glycosylation with age increase the risk of infection and autoimmune disease by causing inflammation. Researchers are focusing on therapeutic strategies that target the glycosylation mechanism⁵.

Defects in glycosylation are also linked to diseases such as cancer, diabetes, and atherosclerosis.

Ubiquitination: Role in protein degradation and cellular cleanup

Ubiquitination is a reversible post-translational modification that primarily targets lysine residues, playing a key role in protein degradation via the ubiquitin-proteasome pathway and regulating processes like cell proliferation, DNA repair, and apoptosis. This PTM occurs in the cytoplasm, ER, nucleus, Golgi apparatus, and mitochondrial matrix.

Key steps in the ubiquitination process

An E1 enzyme activates ubiquitin by forming a high-energy thioester bond between its C-terminal glycine and a cysteine residue on the E1 enzyme, which requires ATP hydrolysis.
The activated ubiquitin is subsequently transported to a cysteine residue on an E2 enzyme to form a thioester bond.
The E3 ubiquitin ligase is responsible for detecting the target protein (substrate) and allowing ubiquitin transfer from the E2 to a lysine residue on the substrate. Different E3 ligases have distinct substrate specificities, which allows for precise control over protein degradation⁶.

Variants of ubiquitination

Ubiquitination generally changes based on the coupling of ubiquitin molecules within a polyubiquitin chain, with the most obvious differences being

• Monoubiquitination: The addition of a single ubiquitin molecule to a substrate, which often regulates protein localization, DNA repair, or receptor endocytosis.

• Multi-monoubiquitination: When numerous lysine residues on the substrate are ubiquitinated, each with a single ubiquitin molecule, and this can impact protein transport and signaling⁷.

• Polyubiquitination: Different lengths of chains of ubiquitin molecules and the substrate, with the length of the chain exerting different effects:

  • K48-linked chains cause proteosomes to degrade proteins.
  • K63-linked chains help to transmit signals, activate kinases, and respond to DNA damage.
  • K11-linked chains control the cell cycle.
  • K29/K33-linked chains are less studied but implicated in immune signaling and protein stability⁸.

Key pathways controlled by ubiquitination include:

• Cell cycle regulation: Degrades proteins like cyclins and cyclin-dependent kinases (CDKs) to ensure normal cell cycle progression.
• Signaling modulation: Alters the localization of signaling proteins or targets them for destruction.
• DNA repair: Ubiquitinates proteins involved in DNA repair to aid in recognition and repair processes.
• Endocytic pathway: Marks proteins for degradation via the endocytic pathway.
• Immune regulation: Degrades receptors and cytokine components to regulate immunological signaling.
• Autophagy: Directs proteins to the autophagosome for degradation.
• Gene expression control: Influences transcription factor stability and activity, impacting gene expression.

Consequences of aberrant ubiquitination

Dysregulated ubiquitination impacts multiple cellular processes, contributing to various diseases:

• Neurodegenerative disorders: Impaired proteasomal degradation leads to toxic protein aggregation, as seen in:

  • Parkinson’s disease - Accumulation of α-synuclein
  • Alzheimer’s disease - Aggregation of tau and amyloid-β
  • Huntington’s disease - Mutant huntingtin protein accumulation9

• Cancer: Disrupted ubiquitin signaling promotes tumorigenesis through:

  • Loss of tumor suppressors – for example, p53 degradation by MDM2
  • Overactive ubiquitin ligases – for example, Skp2-mediated cell cycle dysregulation10

• Autoimmune diseases: Aberrant ubiquitination in NF-κB signaling is linked to conditions such as:

  • Rheumatoid arthritis
  • Lupus

• Aging and cellular senescence: Dysfunctional ubiquitin-proteasome activity results in:

  • Protein accumulation
  • Oxidative stress
  • Cellular failure

Targeted medicines include proteasome inhibitors, Deubiquitinase (DUB) inhibitors, and protein degradation methods. Stability and activity influence gene expression¹¹.

Methylation

Methylation occurs largely in the nucleus and cytoplasm, depending on the target protein and its function, and it is a reversible post-translational modification that regulates protein function, location, and interaction. It transfers a methyl group (-CH₃) from S-adenosylmethionine (SAM) to amino acids like lysine and arginine. Methyltransferases catalyze the process, whereas demethylases remove methyl groups, resulting in reversible and dynamic methylation.

This PTM plays a critical role in various cellular functions, including:

• Histone methylation: Modulates chromatin structure and gene expression. For instance, H3K4me3 promotes transcription, while H3K27me3 represses gene activity. DNA methylation, mediated by DNMTs, works in conjunction with histone modifications to establish long-term gene regulation¹².
• Lysine methylation: Lysine residues can undergo mono-, di-, or tri-methylation, influencing chromatin remodeling and transcription. This process is mediated by SET domain methyltransferases (eg, SUV39H1, EZH2) and reversed by Jumonji-domain demethylases (eg, JMJD3)¹³.
• Arginine methylation: Mono- or di-methylation (symmetric or asymmetric) of arginine residues regulate RNA processing and signaling. Protein arginine methyltransferases (PRMTs), such as PRMT1 and PRMT5, are key enzymes involved in this modification¹³.
• Methylation of other amino acids: Histidine, glutamine, and asparagine methylation, though less common, are essential for bacterial signaling and enzyme regulation.
• RNA-binding protein regulation: Arginine methylation modulates RNA-binding proteins, impacting mRNA splicing, translation efficiency, and stability¹⁴.
• Non-histone protein methylation: Extends beyond chromatin regulation to influence critical physiological processes, including cell cycle progression, immune responses, stress tolerance, and the function of key signaling proteins such as p53, NF-κB, and STAT3¹⁵.

The mechanism of methylation

The methylation process begins with the identification of a certain amino acid residue (often arginine or lysine) that needs to be methylated by a particular methyltransferase enzyme of the target protein. A methyl donor molecule, like S-adenosyl methionine (SAM), serves as the source of the methyl group and is bound by the methyltransferase. The methyltransferase catalyzes the transfer of the methyl group from SAM to the desired amino acid residue on the protein, resulting in a covalent link.

Following methylation, the modified protein is released, and the enzyme can proceed to further methylate target proteins. Dysregulation of methylation is linked to diseases such as cancer, diabetes, and neurological disorders¹⁶.

For example, the overactivity of EZH2, a histone methyltransferase found in several malignancies, induces abnormal gene silencing, which drives carcinogenesis. Methylation of p53 reduces tumor suppression, impairing growth control. Additionally, hypermethylation of CpG islands in gene promoters silences tumor suppressors, such as MLH1, in colorectal cancer, allowing for uncontrolled growth. These epigenetic changes play significant roles in carcinogenesis and progression.

The polycomb repressive complex 2 member EZH2 catalyzes the trimethylation of H3K27me3, a change known to be a component of the histone code. According to the histone code theory, methylation, acetylation, and ubiquitination are among the chemical changes of histone proteins thought to exhibit unique functions in the epigenetic control of gene transcription. Long-term transcriptional suppression linked to EZH2-mediated catalysis of H3K27me3 contributes to carcinogenesis.

Alzheimer’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis are all associated with altered methylation of key proteins, such as tau, TDP-43, and α-synuclein. Moreover, PRMT dysregulation reduces neuronal plasticity, which leads to neurodegeneration and a reduction in cognitive function.

DNA hypomethylation causes genomic instability and raises vulnerability to age-related illnesses, whereas hypermethylation of mitochondrial genes hinders cellular energy metabolism. Changes in histone methylation speed up the tissue deterioration and functional loss associated with aging by impeding stem cell regeneration¹⁷.

Chronic inflammation has been associated with abnormal patterns of methylation, which have been connected to the onset and progression of autoimmune disorders such as systemic lupus erythematosus and rheumatoid arthritis. Excessive inflammation can result from the deregulation of these signaling pathways, which are essential for controlling inflammatory responses and are caused by aberrant methylation.

Acetylation

Acetylation, a reversible modification, is catalyzed by acetyltransferases and deacetylases. It primarily targets lysine residues by adding acetyl group (COCH₃) and plays an important role in processes like:

• Chromatin stability
• Cell cycle control
• Metabolism

There are two primary types of acetylation:

• N-terminal acetylation: The addition of an acetyl group to a protein’s amino terminus regulates stability, localization, and interactions. It can signal degradation or protect proteins. Dysregulation is linked to developmental delays, cognitive impairments, and other pathologies. This modification is among the most common in eukaryotic proteins.
• Lysine acetylation, the reversible addition of an acetyl group to the ε-amino group of a lysine residue, regulates gene expression, protein interactions, and metabolism. It is crucial for cellular homeostasis, and its dysregulation is linked to various diseases¹⁸.

Acetylation influences several critical cellular pathways. Histone acetylation promotes transcriptional activity and results in a more flexible chromatin structure, particularly by modifying the charge of lysine residues and regulating processes like gene expression through histone modification. Conversely, deacetylation may result in chromatin condensation and transcriptional inhibition¹⁹.

Protein acetylation is a crucial PTM that profoundly regulates the action of several metabolic enzymes. This directly affects the “metabolic flux,” or movement of metabolites through metabolic pathways, inside a cell. This PTM occurs in multiple compartments within a cell, including the cytosol, nucleus, mitochondria, and the lumen of the ER²⁰.

Acetylation can affect how quickly proteins break down and where they are found inside cells, which can change how they function in the cell.

This image is from an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Reference: Impact of N-Linked Glycosylation on Therapeutic Proteins. Baoquan Chen, Wenqiang Liu, Yaohao Li, Bo Ma, Shiying Shang, Zhongping Tan. Molecules 2022, 27(24), 8859; https://doi.org/10.3390/molecules27248859

Figure 2: Some examples of post-translational modifications. Gal: galactose; Man: mannose; GlcNAc: N-acetylglucosamine; Neu5Ac: sialic acid; Fuc: fucose.

SUMOylation

SUMOylation involves the attachment of small ubiquitin-related modifier (SUMO) proteins to lysine residues and regulates essential cellular processes such as transcription, chromatin organization, and genome integrity. Although SUMOylation predominantly occurs in the nucleus, under certain circumstances (like stress), it can also occur in the cytoplasm. This PTM is extremely reversible and closely controlled by SUMO-specific proteases (SENPs), resulting in dynamic protein regulation. SENP1 and SENP2 remove SUMO modifications, restoring protein function and ensuring homeostasis. This reversibility affects SUMOylation levels in response to cellular stress and physiological changes.

A cascade of enzymes mediates the process:

• E1 Activating Enzyme: Activates SUMO.
• E2 Conjugating Enzyme: Transfers SUMO to the target protein.
• E3 Ligase: Facilitates the transfer of SUMO from E2 to the substrate²¹.

The roles of SUMO paralogs (SUMO1-4) vary. SUMO1 regulates gene expression by modulating chromatin structure and nuclear transport. SUMO2/3 uses SUMO-targeted ubiquitin ligases to control stress responses, proteostasis, and degradation through polySUMO chains. Fewer studies have been conducted on SUMO4, which is associated with inflammatory diseases and immunological regulation.

Variants of SUMOylation

Monomeric SUMOylation alters a lysine residue, influencing protein interactions and location, including p53 control. PolySUMOylation by SUMO2/3 generates chains required for breakdown, stress response, and proteostasis, and SUMOylation is linked to the ubiquitin-proteasome system via SUMO-targeted ubiquitin ligase²².

SUMOylation also works in concert with other PTMs. It competes with ubiquitination for lysine residues, affecting protein stability. For example, SUMOylation of IκBα occurs on the Lys21 residue, which is also a ubiquitination target, thus protecting IκBα from degradation. Phosphorylation-dependent SUMOylation promotes SUMOylation in transcription factors such as c-Jun, whereas SUMOylation frequently inhibits transcription in contrast to acetylation²³.

Diseases and aging-related processes associated with SUMOylation

• Dysregulation of SUMOylation can impair the normal clearance of damaged proteins and cause misfolded proteins to accumulate and form toxic aggregates, which are the hallmarks of neurodegenerative disorders.
• Aberrant SUMOylation contributes to Alzheimer’s, Parkinson’s, and Huntington’s disease by altering tau, α-synuclein, and huntingtin aggregation, compromising proteostasis, and disturbing mitochondrial function, leading to neurodegeneration²⁴.
• SUMOylation regulates ion channels and stress responses. Dysregulation results in heart failure and injury from ischemia and impairs myocardial survival and repair.
• SUMOylation influences transcription, the cell cycle, and DNA repair. Hyper-SUMOylation inhibits p53, which in turn promotes tumor development and resistance to apoptosis, ultimately causing cancer progression and therapeutic resistance.
• Age-related reductions in SUMOylation affect genomic stability and stress response. Impaired SUMOylation accelerates aging-related diseases, cellular senescence, and DNA damage. Recent research has connected SUMOylation of a specific RNA binding protein (CAR-1) in the germline to organismal aging²⁵.

Lipidation: impact on biological processes

Lipidation, a post-translational modification involving the covalent attachment of lipids like palmitic acid to proteins, plays an important role in processes such as protein function regulation, signal transduction, and neuronal development. There are three types of lipidation:

• Palmitoylation: Addition of palmitic acid, a 16-carbon saturated fatty acid, to cysteine residues in proteins via thioester bonds (S-palmitoylation) or amide linkages (N-palmitoylation). This reversible change, mediated by palmitoyl transferases and acyl-protein thioesterases, enables dynamic regulation of membrane-associated proteins. It is necessary for synaptic plasticity, receptor trafficking, and immune signaling.
• Myristoylation: Addition of a myristoyl group, a C14-saturated carboxylic acid, to the N-terminal end of glycine residues. This irreversible change, which enhances membrane targeting and protein-protein interactions, is catalyzed by N-myristoyltransferases. Myristoylation is essential for signal transduction pathways, including G-protein and kinase activation.
• Prenylation: the covalent attachment of hydrophobic isoprenoid groups (farnesyl or geranylgeranyl) to proteins, specifically cysteine residues, via CaaX motifs. Prenylation, which is controlled by geranylgeranyl transferase and farnesyltransferase and controls intracellular trafficking, cytoskeletal dynamics, and cell proliferation, is necessary for the activation of Ras, Rho, and Rab GTPase.

Proteins undergo prenylation and myristoylation in the cytoplasm prior to proteins reaching their target membranes, and palmitoylation on the cytoplasmic face of membranes, including the plasma membrane and Golgi apparatus. Dysregulated lipidation has been linked to several diseases, such as Alzheimer’s, schizophrenia, Huntington’s disease, and various cancers.

Key synaptic proteins (such as PSD-95 and SNAP-25) have impaired S-palmitoylation, which impairs synaptic function and is linked to Huntington's disease, schizophrenia, and Alzheimer's disease²⁶.

Neuronal survival is also hampered by defective myristoylation²⁷. Oncogenic Ras and Rho GTPases are hyperprenylated, which promotes tumorigenesis by increasing invasion and proliferation. Anticancer treatments using FTase inhibitors have been investigated²⁸.

Cardiomyopathy and ischemic heart disease are exacerbated by aberrant protein myristoylation, which affects calcium channel control and heart muscle contraction.

Defects in lipidation compromise mitochondrial activity, metabolic signaling, and membrane integrity, hastening immunological and cognitive deterioration associated with aging²⁹.

Nitrosylation: Role in a broad spectrum of proteins

Nitrosylation is a reversible post-translational modification where nitric oxide (NO) covalently binds to cysteine thiolsto form S-nitrosothiols, regulating over 3,000 proteins involved in processes like DNA repair, transcription, and apoptosis. This redox-based modification plays a crucial role in modulating protein stability, localization, function, and interactions.

Dysregulation of S-nitrosylation (SNOs) that influences a wide array of cellular functions through its effects on protein structure and activity contributes to various diseases by affecting protein stability, interactions, and other modifications. S-nitrosylation takes place in the cytosol, mitochondria, nucleus, endoplasmic reticulum (ER), and chloroplasts in plants.

Mechanism and pathways of S-Nitrosylation

Nitric oxide synthases (NOSs), which generate NO from L-arginine, regulate S-nitrosylation. The three major NOS isoforms that contribute to this modification are neuronal NOS, which is involved in memory, synaptic plasticity, and neurotransmission; endothelial NOS, which controls vasodilation, blood flow, and cardiovascular homeostasis; and inducible NOS, which produces NO in response to immune activation and inflammation, as well as in pathogen defense and cellular stress responses³⁰.

Denitrosylases, such as S-nitrosoglutathione reductase and thioredoxin, can reverse S-nitrosylation either enzymatically or spontaneously. The equilibrium between S-nitrosylation and denitrosylation ensures proper cellular function and oxidative stress response. S-nitrosylation is the addition of NO to cysteine residues, which alters protein activity, stability, and interactions; for example, SNO of p53 regulates apoptosis in response to DNA damage³¹.

• Metal nitrosylation impacts oxygen transport and respiration and involves NO coordination with metal centers in metalloproteins, including hemoglobin and cytochrome C oxidase³². Neurodegenerative diseases such as Parkinson's disease are exacerbated by abnormal S-nitrosylation of parkin and UCH-L1, causing a reduction in ubiquitin-proteasome activity³³. By facilitating protein aggregation and oxidative stress, SNO of tau and DJ-1 is associated with Huntington's and Alzheimer's diseases³⁴.
• Dysregulated eNOS-mediated SNO contributes to hypertension, atherosclerosis, and heart failure due to compromised vasodilation, endothelial function, and calcium management³⁵.
• The S-nitrosylation of tumor suppressors (eg, PTEN) and DNA repair enzymes promotes uncontrolled cell proliferation and genomic instability³⁶.
• Nitrosative stress causes mitochondrial dysfunction, poor insulin signaling, and inflammation, which hastens aging and age-related disorders like type 2 diabetes and dementia³⁶.

Although it has historically been thought of as a non-enzymatic post-translational modification, research also indicates that the enzymes that catalyze S-nitrosylation or denitrosylation can also control protein S-nitrosylation, which is the covalent attachment of nitric oxide (NO) to cysteine residues. For example, in Escherichia coli, de novo endogenous protein S-nitrosylation is primarily enzymatic.

Glycation

In glycation, reducing sugars react with amino groups in proteins, lipids, or nucleic acids to produce advanced glycation end products (AGEs). Glycation occurs mostly in the extracellular matrix but also in the cytoplasm and mitochondria, particularly under high glucose exposure. This takes place in three stages:

• Schiff base formation is an initial reversible reaction where a sugar molecule directly attaches to a protein's amino group, forming a Schiff base.
• Amadori rearrangement is a more stable rearrangement of the Schiff base and is considered the primary early glycation product.
• AGE buildup is a complex mixture of highly reactive, cross-linked molecules formed through further rearrangements and oxidation of early glycation end products, including carboxymethyl-lysine, methylglyoxal-derived AGEs, etc³⁷.

Interaction of AGEs with their receptors leads to oxidative damage and inflammation. The glyoxalase system detoxifies reactive intermediates. However, excessive glycation leads to diabetes, aging, and neurological diseases such as Parkinson's and Alzheimer's disease. Glycation accelerates the onset of diabetes and affects cellular homeostasis through vascular damage, neuropathy, and retinopathy; it promotes protein aggregation and oxidative damage in neurodegenerative diseases³⁷.

Carbonylation

While cells use antioxidant defense mechanisms such as superoxide dismutase and catalase to prevent oxidative damage, prolonged oxidative stress causes uncontrolled carbonylation. This PTM occurs primarily in mitochondria. Unlike reversible PTMs like phosphorylation or ubiquitination, carbonylation is an irreversible, non-enzymatic PTM in which carbonyl groups (ketones or aldehydes) are added to protein side chains, particularly targeting lysine, arginine, proline, and threonine residues through reactions carried out by reactive oxygen species or reactive carbonyl species³⁸.

Carbonylation is irreversible and marks proteins for destruction, leading to their loss of function and aggregation; their accumulation is a hallmark of oxidative stress-related diseases, making carbonylation an important biomarker for aging and disease. Carbonylation plays a crucial role in aging and age-related diseases, as oxidized proteins accumulate due to declining proteasomal and autophagic clearance.

• In neurodegenerative diseases like Alzheimer’s and Parkinson’s, excessive carbonylation affects key proteins, promoting aggregation and neuronal dysfunction.
• In diabetes, oxidative stress causes carbonylation, which leads to β-cell destruction and insulin resistance³⁹.
• Cardiovascular disorders are also associated with inflammation and carbonylation-induced endothelial dysfunction.

Lysine glutarylation

For example, lysine glutarylation (Kglu), is a specific form of protein modification where a five-carbon glutaryl group is attached to a lysine amino acid on a protein. A particular “eraser” enzyme can reverse Kglu, removing the glutaryl group and enabling dynamic control of protein activity. Research is being done to learn more about its functions in controlling cellular functions.

Proteolytic cleavage and other post-translational modifications

Proteolytic cleavage, along with other post-translational modifications like lysine acetylation and ubiquitination, precisely alters protein structure and function, enabling the regulation of complex processes such as enzyme activation, protein turnover, and cellular response to stress.

Proteolytic cleavage is a type of PTM where specific peptide bonds in a protein are cleaved by proteases, leading to the activation or inactivation of the protein. This process plays a vital role in:

• Regulating protein function.
• Controlling cellular processes.
• Modulating signaling pathways by removing or rearranging protein fragments.

Disulfide bond formation

Disulfide bond formation is an important post-translational modification that stabilizes protein structure and regulates protein folding, function, and interactions. The bond formation occurs when two cysteine residues in a protein form a covalent bond between their thiol (-SH) groups, resulting in a -S-S- linkage.

It can occur both intracellularly and extracellularly through various enzymatic and non-enzymatic mechanisms, affecting processes like protein stability, enzyme activity, and immune responses.

ADP ribosylation: role in cellular processes

ADP-ribosylation is a process in which ADP-ribose molecules are covalently attached to proteins, nucleic acids, and other biomolecules, influencing various cellular processes such as DNA repair, transcription, and cell signaling. Understanding the mechanisms and implications of this modification offers valuable insights into its potential as a therapeutic target for various diseases, particularly those involving dysregulated cellular processes, including cancer treatment, neurodegenerative diseases, and inflammation.

The role of the endoplasmic reticulum, Golgi apparatus, and other compartments

The ER promotes folding, N-linked glycosylation, lipidation, and disulfide bond formation and is thus necessary for PTMs in membrane proteins and secretory proteins. The quality control carried out by ER, which includes ER-associated degradation and the unfolded protein response - when unfolded proteins accumulate in the ER, leading to upregulation of chaperones and protein folding machinery to alleviate stress - maintains homeostasis by eliminating misfolded proteins or refolding.

ER-phagy, a process in which autophagosomes engulf and degrade damaged ER components, helps reduce two key factors in neurodegenerative disorders: cellular stress and protein aggregation.

The Golgi apparatus is important for PTMs, ensuring protein maturation, including vesicle trafficking and the modification of macromolecules, and plays a vital role in cellular stress responses and unconventional protein secretion pathways. It mediates O-linked glycosylation (adding sugars to serine/threonine hydroxyl groups), sulfation of tyrosines/carbohydrates (essential for interactions and extracellular matrix organization), and phosphorylation of oligosaccharides on lysosomal proteins (guiding their cellular destination)⁴¹.

Defects in Golgi function are linked to various diseases and help understand disease pathogenesis and potential therapeutic strategies.  Further, disruptions in these PTMs contribute to diseases like congenital disorders of glycosylation and cancer metastasis, suggesting the key role of Golgi bodies in cellular homeostasis and disease pathogenesis.

Other cell compartments, such as lysosomes and peroxisomes, contribute to post-translational modifications by degrading and recycling proteins, including removing certain PTMs.

Influence on protein localization and interactions

PTMs like glycosylation and phosphorylation determine protein localization and modulate protein-protein interactions, with phosphorylated proteins in central network positions and glycosylated proteins at the periphery.

Significance of post-translational modifications in biological processes

Post-translational modifications alter protein properties such as charge, hydrophobicity, and conformation, influencing their function, interactions, and aggregation. Any disruption in modifications can alter biological processes and lead to cancer.

Effect on pharmacokinetics and pharmacodynamics properties of proteins

PTMs affect stability and half-life, clearance, binding to transport protein, and bioavailability, thereby impacting pharmacokinetic and pharmacodynamic properties of proteins especially ones to be used for therapeutic applications.

For instance, ubiquitination targets proteins for proteasomal degradation, impacting drug stability and bioavailability¹¹, and acetylation and SUMOylation regulate protein-protein interactions and transport, affecting therapeutic efficacy⁴².  Phosphorylation modulates receptor binding and activation, altering drug potency. Disulfide bond formation stabilizes monoclonal antibodies and other biologics, ensuring prolonged activity⁴³.

Furthermore, by binding oligosaccharides to asparagine residues, N-glycosylation improves protein stability, solubility, and resistance to proteolysis, extending its half-life in circulation. N-glycosylation influences immune recognition and decreases unpleasant effects by modifying immunogenicity. By driving protein structure and characteristics, glycosylation ensures optimal ER function. N-glycosylation management is critical to the stability, safety, and efficacy of therapeutic proteins⁴⁴.

Linking post-translational modification abnormalities to disease pathogenesis

Abnormalities in PTMs can disrupt cellular processes, leading to the dysregulation of key pathways and contributing to the development of various diseases, including cancer, neurodegenerative disorders, and metabolic diseases.

Cancer

The dysregulation of PTMs is implicated in cancer progression, making a key focus for understanding protein functions and potential therapeutic targets.

Dysregulated phosphorylation in cancer activates oncogenes or deactivates tumor suppressors, resulting in uncontrolled proliferation. Overactive receptor tyrosine kinases (RTKs) cause cancer. Therefore, kinase inhibitors such as imatinib, which target BCR-ABL in chronic myeloid leukemia, are effective therapeutic methods.

Acetylation alters the regulation of genes involved in the cell cycle and apoptosis. Histone hypoacetylation inhibits tumor suppressors. HDAC inhibitors restore acetylation, reactivating repressed genes while decreasing tumor development, making them powerful anticancer medicines.

This image is from an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Reference: The role of protein acetylation in carcinogenesis and targeted drug discovery. Front. Endocrinol. 12 September 2022. Sec. Cancer Endocrinology. Volume 13 - 2022  https://doi.org/10.3389/fendo.2022.972312

Figure 3: The role of protein acetylation in carcinogenesis and targeted drug discovery

Aberrant glycosylation in cancer promotes tumor growth and immune evasion. Altered glycans promote cell dissociation and invasion, hence increasing metastasis. These changes aid malignancies in evading immune identification. Understanding glycosylation alterations has resulted in glycan-based biomarkers and treatment targets in oncology.

Altered ubiquitination either destroys tumor suppressors or stabilizes oncogenic proteins. Dysregulated SUMOylation impacts transcription factors and DNA repair proteins, which promote genomic instability and tumor growth. Bortezomib, a proteasome inhibitor, disrupts these processes and serves as a cancer therapy.

PD-L1, an immune checkpoint protein, is modified by phosphorylation, glycosylation, acetylation, and ubiquitination, influencing its stability, localization, and immune evasion. Similarly, p53 experiences a variety of PTMs, including phosphorylation, acetylation, and SUMOylation, which control its activation in response to stress.

This image is from an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Reference: (P53 modifications: Exquisite decorations of the powerful guardian. July 2019. Journal of Molecular Cell Biology 11(7). DOI:10.1093/jmcb/mjz060

Figure 4: Post-translational changes can control p53 in many ways. (A) p53 phosphorylation at S33, T81, and S315 creates a docking motif for peptidyl-prolyl cis-trans isomerase (Pin1). (B) Phosphorylation at the N-terminus of p53 provides negative electrostatic forces, allowing CREB-1 binding protein (CBP)/p300 to interact with positive electrostatic forces. (C and D) Mdm2 (mouse double minute 2) polyubiquitinates p53 for proteasomal destruction but monoubiquitinates it for nuclear export. (E) Tip60's acetylation at K120 may enable p53 to shift conformation, allowing it to attach to a specific target gene promoter. (F) Specific p53 alterations may aid in phase separation with other regulators.

Neurogenerative diseases

Abnormal PTMs cause protein misfolding and cytotoxicity, particularly in neurodegenerative disorders.

Tau protein, which is required for microtubule integrity, collects into neurofibrillary tangles in Alzheimer's disease due to excessive phosphorylation at certain sites, affecting neuronal structure and function.  These tangles disrupt the cytoskeleton of neurons, preventing axonal transit and increasing the risk of neuronal death.

Parkin, a protein involved in mitophagy—the process of removing damaged mitochondria—becomes faulty in Parkinson's disease due to abnormal phosphorylation. Impaired mitophagy causes damaged mitochondria to accumulate, resulting in hazardous reactive oxygen species (ROS) that destroy neurons and accelerate the disease.

In Huntington's disease, PTMs like acetylation, SUMOylation, ubiquitination, and palmitoylation are implicated, with phosphorylation being the most extensively studied and potentially critical for disease progression.

The primary PTM linked to the progression of Huntington's disease is phosphorylation of the huntingtin protein, particularly at certain serine and threonine residues, because it affects the protein's aggregation propensity and cellular localization, changes subcellular localization, and impairs cellular signaling pathways, all of which lead to neuronal damage.

Autoimmune diseases

PTMs in self-proteins, such as citrullination and acetylation, can trigger autoimmune responses by creating neoepitopes that break immune tolerance, which can lead to conditions like:
The conversion of the amino acid arginine to citrulline, a vital step in the onset of RA, is one of the best-studied examples of citrullination.

Additional PTMs such as phosphorylation, glycosylation, acetylation, and oxidation may also play a role in the onset of autoimmune disorders. Specific autoimmune disorders related to PTMs:

• A wide spectrum of PTMs on diverse proteins, including changes that influence protein stability and antigen presentation, have been linked to systemic lupus erythematosus.
• PTMs on pancreatic beta cell proteins, such as insulin deamidation, can activate an autoimmune response against beta cells, resulting in insulin shortage and, ultimately, type 1 diabetes.
• Phosphorylation of the cholinergic receptor is associated with the formation of autoantibodies against this receptor, which causes muscular weakness known as myasthenia gravis.

Moreover, environmental factors, like bacterial and viral infections, as well as genetic predispositions, further exacerbate the formation of PTMs, contributing to the development and progression of autoimmune diseases.

Methods for studying post-translational modifications

Detection methods for studying PTMs help in understanding the protein function and stability. These techniques include mass spectrometry, western blotting, immunoprecipitation, and enzyme-linked immunosorbent assays (ELISA) to identify and analyze specific protein modifications. Advanced methods such as protein/antibody arrays, surface plasmon resonance, and Bi-layer interferometry are gaining popularity in detecting and analyzing PTMs in proteins. Techniques for PTM detection and analysis enable the identification, quantification, and characterization of PTMs in proteins. A few of these methods include:

Mass spectrometry (MS)

MS enables sensitive and specific detection of PTMs, identifying modification sites and proteins with enhanced accuracy using techniques like stable isotope labeling, data-independent acquisition (DIA), and enrichment methods. It also plays an important role in detecting and quantifying PTMs by providing the mass accuracy and resolving power needed to identify novel and pathological PTMs.

ELISA

• Antibody-based methods, relying on high-quality, modification-specific, or recombinant antibodies, are widely used for detecting and quantifying PTMs like phosphorylation due to their specificity, ease of use, and improved detection affinity.
• Protein arrays are powerful tools for studying PTMs, enabling high-throughput analysis of protein interactions. By using specific antibodies or binding molecules such as lectins, these arrays provide a snapshot of PTM patterns, offering insights into cellular signaling, disease mechanisms, and biomarker discovery with precision and scalability.

Chromatography

Chromatographic techniques, particularly liquid chromatography, are used to separate, enrich, and purify post-translationally modified proteins or peptides from complex samples, often complementing mass spectrometry for detailed PTM analysis.

Chromatographic methods are also important for profiling primary antibody and secondary antibody structure, post-translational modifications, and quality to meet regulatory standards for therapeutic applications.  Recent advances in resin chemistry have evolved high-resolution techniques such as high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and hydrophilic interaction liquid chromatography (HILIC) for characterizing and elucidating the structure of carbohydrates, particularly monosaccharides, oligosaccharides, and glycans. These are powerful techniques for ensuring product quality in biopharmaceutical development.

Electrophoretic techniques

Proteins are separated according to size and charge using SDS-PAGE and 2D electrophoresis, which reveals migratory shifts caused by PTM. Western blotting with specific antibodies verifies the modifications. Capillary electrophoresis provides a higher separation efficiency for precise PTM analysis. Following gel electrophoresis, mobility shifts can be recognized by western blotting, which is useful for undetected changes like phosphorylation shifts or ubiquitin ladders.

Surface-based methods

Surface plasmon resonance and Bi-layer interferometry are label-free technologies used for studying protein interaction and PTMs. These techniques enable real-time detection of biomolecular interactions by measuring binding kinetics, affinity, and specificity.

Advanced techniques

Recent advances in techniques such as nanopore-based detection, NMR spectroscopy, and cryo-electron microscopy (Cryo-EM) provide detailed insight into protein structure with respect to the complexity of post-translational modifications. Nanopore facilitates single-molecule resolution for detecting PTMs by analyzing changes in ionic current as proteins pass through nanopores.

NMR spectroscopy helps establish PTMs and their structural impact, such as phosphorylation-induced conformational changes, and Cryo-EM visualizes PTMs and their effects on macromolecular complexes at near-atomic resolution. Furthermore, human-induced pluripotent stem cell models have significantly enhanced the understanding of PTMs, such as those of tau protein, including aggregation patterns, and their role in neurodegenerative diseases, offering insights for therapeutic interventions.

Challenges and the future of research on PTMs

Research on PTMs is challenging because of their vast diversity, different chemical properties, and biological activities, potentially resulting in significantly heterogeneous gene products. Understanding PTMs requires the integration of biology, chemistry, and computational approaches.

PTMs are transient and reversible, making it challenging to detect spatiotemporal protein activity modulation. This necessitates advanced approaches like computational modeling and high-resolution mass spectrometry.

PTM research also faces specificity issues due to low abundance, instability, and site-specific detection hurdles, requiring precise analytical methods and technically difficult, resource-intensive enrichment approaches⁴⁵.

Future research in post-translational modifications should focus on developing more advanced detection techniques, enhancing our understanding of their roles in diseases, and translating PTM analysis into clinical diagnostics and therapeutic interventions.

Genetic code expansion (GCE) technology, which uses orthogonal aminoacyl-tRNA synthetase/tRNACUA pairs to incorporate unnatural amino acids into proteins, allows the site-specific addition of unnatural amino acids, resulting in uniform protein creation with precise PTMs. GCE can be used in both prokaryotic and eukaryotic cells⁴⁶.

The integration of bioinformatics tools, AI, and machine learning in PTM research has improved the prediction and identification of PTM sites, offering faster and more accurate solutions compared to traditional methods. Advanced mass spectrometry techniques and machine learning-based analytics further enhance data analysis, aiding in the understanding of cellular mechanisms and disease progression.

Single-cell analysis has revolutionized PTM research by enabling the study of cellular heterogeneity and rare cell characterization through advanced microfluidics, imaging, and omics techniques.

Recent innovations in bioinformatics, including trajectory inference methods and machine learning algorithms, have improved the resolution of cellular dynamics and the integration of single-cell data. However, challenges such as noisy data and technical barriers highlight the need for refined computational tools to enhance disease diagnostics and drug development.

Emerging PTM types and their implications in molecular biology

Recent developments in post-translational modifications include newly identified acylation types such as propionylation, succinylation, and lactylation, which regulate signal transduction, metabolism, and other biological processes.

Less well-studied PTMs, including acylation, lipid-related, and metabolite-related modifications, play important roles in tumorigenesis and cancer development by influencing growth, metabolism, immune response, and mechanical stress in the tumor microenvironment (TME).

Comprehensive PTM analysis holds promise for identifying therapeutic targets, advancing tumor immunotherapy, and improving clinical outcomes by addressing immunosuppression within the TME.

Potential in personalized medicine and proteomics

PTMs are essential in regulating physiological processes such as signal transduction, metabolism, and protein turnover. This makes them central to diseases like cancer and age-related pathologies. Profiling PTMs can reveal disease-specific patterns, enabling targeted therapeutic interventions and advancing personalized medicine.

Interdisciplinary approaches and advancements in PTM analysis

Recent advancements in the analysis of PTMs have emerged from interdisciplinary approaches that integrate various fields, including proteomics, bioinformatics, and machine learning. Interdisciplinary approaches in PTM analysis are advancing cancer immunotherapy by exploring immune-related PTMs and developing inhibitors, such as phosphorylation inhibitors and adoptive cell therapies.

These methods aim to improve tumor immunity through targeted interventions, including knocking out PTM-related proteins to enhance therapeutic outcomes.

Machine learning techniques are increasingly used to predict PTM sites, analyze relationships between PTMs and diseases, and address data imbalance issues in datasets. These methods enhance the accuracy of predictions related to protein folding, activity, and function.

Further, the use of data-independent acquisition (DIA) mass spectrometry, deep learning, computational tools, and improved enrichment techniques have enhanced the accuracy, depth, and scale of PTM analysis.

FAQs

How do PTMs contribute to the complexity of organisms?

PTMs add complexity to organisms by expanding protein function beyond genetic encoding. They allow for the regulation of various cell processes without altering the DNA. They govern signaling, gene expression, and metabolism, among many other processes, allowing for fast cellular responses. PTMs facilitate protein interactions, promote tissue specialization, and regulate chromatin dynamics. This additional regulatory layer increases adaptability and functional diversity.

What role do PTMs play in disease development?

Numerous PTMs play a significant role in the development of several diseases; dysregulation of PTMs can alter the activity and function of proteins, interfering with essential cellular pathways, leading to aberrant cell behavior and ultimately hastening the onset of various illnesses, such as cancer, neurological conditions, and metabolic diseases. For example, improper phosphorylation or misfolding of proteins can contribute to diseases like cancer, Alzheimer's disease, and diabetes.

How are PTMs analyzed and detected in modern biology?

PTMs are analyzed and detected using techniques like mass spectrometry, which helps identify and quantify modifications, and western blotting, which detects specific modified proteins. Other methods, such as enzyme-linked immunosorbent assays (ELISA) and protein microarrays, are also used to study PTMs in more detail. Additionally, machine learning and AI are being increasingly employed to analyze large-scale data and predict PTM patterns, enhancing the accuracy and efficiency of PTM studies.

References

1. Zhong Q, Xiao X, Qiu Y, et al. Protein posttranslational modifications in health and diseases: Functions, regulatory mechanisms, and therapeutic implications. MedComm. (2020).
2. Ardito F, Giuliani M, Perrone D, et al. The crucial role of protein phosphorylation in cell signaling and its use as targeted therapy (Review). Int J Mol Med. 40, 271-280 (2017).
3. Hirakawa H, Kurushima J, Hashimoto Y, et al. Progress overview of bacterial two-component regulatory systems as potential targets for antimicrobial chemotherapy. Antibiotics (Basel). 9, (2020).
4. Rawat P, Sehar U, Bisht J, et al. Phosphorylated tau in Alzheimer's disease and other tauopathies. Int J Mol Sci. 23, (2022).
5. He, M., Zhou, X., Wang, X. Glycosylation: mechanisms, biological functions and clinical implications. Sig Transduct Target Ther. 9, 194 (2024).
6. Damgaard, R.B. The ubiquitin system: from cell signaling to disease biology and new therapeutic opportunities. Cell Death Differ. 28, 423–426 (2021).
7. Swatek, K., Komander, D. Ubiquitin modifications. Cell Res. 26, 399–422 (2016).
8. Liu P, Gan W, Su S, et al. K63-linked polyubiquitin chains bind to DNA to facilitate DNA damage repair. Sci Signal. 11 (2018).
9. Schmidt MF, Gan ZY, Komander D, et al. Ubiquitin signaling in neurodegeneration: mechanisms and therapeutic opportunities. Cell Death Differ. 28, (2021).
10. William JNG, Dhar R, Gundamaraju R, et al. SKping cell cycle regulation: role of ubiquitin ligase SKP2 in hematological malignancies. Front Oncol. 14, (2024).
11. Zhao, L., Zhao, J., Zhong, K. et al. Targeted protein degradation: mechanisms, strategies and application. Sig Transduct Target Ther. 7, 113 (2022).
12. Jeong M, Sun D, Luo M, et al. Large conserved domains of low DNA methylation maintained by Dnmt3a. Nat Genet. 46,17-23 (2014).
13. Separovich RJ, Wilkins MR. Ready, SET, Go: Post-translational regulation of the histone lysine methylation network in budding yeast. J Biol Chem. 297, (2021).
14. Ratovitski, T., Kamath, S. V., O'Meally, R. N., et al. Arginine methylation of RNA-binding proteins is impaired in Huntington's disease. Human molecular genetics, 32, 3006–3025 (2023).
15. Lehning, N. A., Morrison, B. E. Nonhistone Lysine Methylation as a Protein Degradation Signal. Journal of chemistry. (2022).
16. Menezo Y, Clement P, Clement A., et al. Methylation: An Ineluctable Biochemical and Physiological Process Essential to the Transmission of Life. Int J Mol Sci. 21, (2020).
17. Liu, R., Zhao, E., Yu, H. et al. Methylation across the central dogma in health and diseases: new therapeutic strategies. Sig Transduct Target Ther. 8, 310 (2023).
18. Christensen, D. G., Xie, X., Basisty, N., et al. Post-translational protein acetylation: an elegant mechanism for bacteria to dynamically regulate metabolic functions. Frontiers in microbiology, 10, 1604 (2019).
19. Chen HP, Zhao YT, Zhao TC. Histone deacetylases and mechanisms of regulation of gene expression. Crit Rev Oncog. 20, 35-47 (2015).
20. Kuhn ML, Rakus JF, Quenet D. Acetylation, ADP-ribosylation and methylation of malate dehydrogenase. Essays Biochem. 68,199-212 (2024).
21. Huang CH, Yang TT, Lin KI. Mechanisms and functions of SUMOylation in health and disease: a review focusing on immune cells. J Biomed Sci. 31, (2024).
22. Wang W, Matunis MJ. Paralogue-Specific Roles of SUMO1 and SUMO2/3 in protein quality control and associated diseases. Cells. 13, (2023).
23. Barbour H, Nkwe NS, Estavoyer B, et al. An inventory of crosstalk between ubiquitination and other post-translational modifications in orchestrating cellular processes. iScience. 26, (2023).
24. Mandel N, Agarwal N. Role of SUMOylation in neurodegenerative diseases. Cells. 11, (2022).
25. Princz A, Pelisch F, Tavernarakis N. SUMO promotes longevity and maintains mitochondrial homeostasis during ageing in Caenorhabditis elegans. Sci Rep. 10, (2020).
26. Peng, J., Liang, D., Zhang, Z. Palmitoylation of synaptic proteins: roles in functional regulation and pathogenesis of neurodegenerative diseases. Cell Mol Biol Lett. 29, 108 (2024).
27. Kanie, T., Ng, R., Abbott, K. L., et al. Myristoylated neuronal calcium sensor-1 captures the ciliary vesicle at distal appendages. bioRxiv : the preprint server for biology. (2023).
28. Karnoub AE, Der CJ. Rho Family GTPases and cellular transformation. In: Madame Curie Bioscience Database [Internet]. Austin (TX): Landes Bioscience; 2000-2013.
29. Nielson JR, Rutter JP. Lipid-mediated signals that regulate mitochondrial biology. J Biol Chem. 293, 7517-7521 (2018).
30. Förstermann U, Sessa WC. Nitric oxide synthases: regulation and function. Eur Heart J. 33, (2012).
31. Xiaomeng Shi, Hongyu Qiu. Post-translational S-nitrosylation of proteins in regulating cardiac oxidative stress. Antioxidants. 9, (2020).
32. Verde C, Giordano D, Bruno S. NO and Heme proteins: Cross-talk between heme and cysteine residues. Antioxidants (Basel). 12, (2023).
33. Nakamura T, Oh CK, Zhang X, Lipton SA. Protein S-nitrosylation and oxidation contribute to protein misfolding in neurodegeneration. Free Radic Biol Med. 172, 562-577 (2021).
34. Choi MS, Nakamura T, Cho SJ, et al. Transnitrosylation from DJ-1 to PTEN attenuates neuronal cell death in Parkinson’s disease models, J. Neurosci. 34, 15123–15131 (2014).
35. Lima B, Forrester MT, Hess DT, et al. S-nitrosylation in cardiovascular signaling. Circ Res. 106, 633-646 (2010).
36. Sharma V, Fernando V, Letson J, et al. S-Nitrosylation in tumor microenvironment. Int J Mol Sci. 22, (2021).
37. Mengstie, M. A., Chekol Abebe, E., Behaile Teklemariam, A., et al. Endogenous advanced glycation end products in the pathogenesis of chronic diabetic complications. Frontiers in molecular biosciences. 9, (2022).
38. Weng, SL., Huang, KY., Kaunang, F.J. et al. Investigation and identification of protein carbonylation sites based on position-specific amino acid composition and physicochemical features. BMC Bioinformatics. 18 (2017).
39. Dinić S, Arambašić Jovanović J, Uskoković A, et al. Oxidative stress-mediated beta cell death and dysfunction as a target for diabetes management. Front Endocrinol (Lausanne). 13, (2022).
40. Wiseman RL, Mesgarzadeh JS, Hendershot LM. Reshaping endoplasmic reticulum quality control through the unfolded protein response. Mol Cell. 82, 1477-1491, (2022).
41. Zhang X. Alterations of Golgi structural proteins and glycosylation defects in cancer. Front cell dev biol. 9, (2021).
42. Lee, J.M., Hammarén, H.M., Savitski, M.M. et al. Control of protein stability by post-translational modifications. Nat Commun. 14, 201 (2023).
43. Ren, T., Tan, Z., Ehamparanathan, V., et al. Antibody disulfide bond reduction and recovery during biopharmaceutical process development-A review. Biotechnology and bioengineering, 118, 2829–2844 (2021).
44. Zhou, Q., Qiu, H. The mechanistic impact of N-glycosylation on stability, pharmacokinetics, and immunogenicity of therapeutic proteins. Journal of pharmaceutical sciences, 108, 1366–1377 (2019).
45. Kitamura N, Galligan JJ. A global view of the human post-translational modification landscape. Biochem J. 480, 1241-1265 (2023).
46. Streit M, Hemberger M, Häfner S, et al. Optimized genetic code expansion technology for time-dependent induction of adhesion GPCR-ligand engagement. Protein Sci. 32, (2023).