Protein methylation

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Protein methylation is a type of post-translational modification featuring the addition of methyl groups to proteins. It can occur on the nitrogen-containing side-chains of arginine and lysine, [1] [2] but also at the amino- and carboxy-termini of a number of different proteins. In biology, methyltransferases catalyze the methylation process, activated primarily by S-adenosylmethionine. Protein methylation has been most studied in histones, where the transfer of methyl groups from S-adenosyl methionine is catalyzed by histone methyltransferases. Histones that are methylated on certain residues can act epigenetically to repress or activate gene expression. [3] [4]

Contents

Methylation by substrate

Multiple sites of proteins can be methylated. For some types of methylation, such as N-terminal methylation and prenylcysteine methylation, additional processing is required, whereas other types of methylation such as arginine methylation and lysine methylation do not require pre-processing.

Arginine

Arginine methylation by type I and II PRMTs Arginine methylation.svg
Arginine methylation by type I and II PRMTs

Arginine can be methylated once (monomethylated arginine) or twice (dimethylated arginine). Methylation of arginine residues is catalyzed by three different classes of protein arginine methyltransferases (PRMTs): Type I PRMTs (PRMT1, PRMT2, PRMT3, PRMT4, PRMT6, and PRMT8) attach two methyl groups to a single terminal nitrogen atom, producing asymmetric dimethylarginine (N G,N G-dimethylarginine). In contrast, type II PRMTs (PRMT5 and PRMT9) catalyze the formation of symmetric dimethylarginine with one methyl group on each terminal nitrogen (symmetric N G,N' G-dimethylarginine). Type I and II PRMTs both generate N G-monomethylarginine intermediates; PRMT7, the only known type III PRMT, produces only monomethylated arginine. [5]

Arginine-methylation usually occurs at glycine and arginine-rich regions referred to as "GAR motifs", [6] which is likely due to the enhanced flexibility of these regions that enables insertion of arginine into the PRMT active site. Nevertheless, PRMTs with non-GAR consensus sequences exist. [5] PRMTs are present in the nucleus as well as in the cytoplasm. In interactions of proteins with nucleic acids, arginine residues are important hydrogen bond donors for the phosphate backbone — many arginine-methylated proteins have been found to interact with DNA or RNA. [6] [7]

Enzymes that facilitate histone acetylation [ citation needed ] as well as histones themselves can be arginine methylated. Arginine methylation affects the interactions between proteins and has been implicated in a variety of cellular processes, including protein trafficking, signal transduction and transcriptional regulation. [6] In epigenetics, arginine methylation of histones H3 and H4 is associated with a more accessible chromatin structure and thus higher levels of transcription. The existence of arginine demethylases that could reverse arginine methylation is controversial. [5]

Lysine

Lysine methylation by PKMTs and demethylation by PKDMs Lysine methylation.svg
Lysine methylation by PKMTs and demethylation by PKDMs

Lysine can be methylated once, twice, or three times by lysine methyltransferases (PKMTs). [8] Most lysine methyltransferases contain an evolutionarily conserved SET domain, which possesses S-adenosylmethionine-dependent methyltransferase activity, but are structurally distinct from other S-adenosylmethionine binding proteins. Lysine methylation plays a central part in how histones interact with proteins. [9] Lysine methylation can be reverted by lysine demethylases (PKDMs). [8]

Different SET domain-containing proteins possess distinct substrate specificities. For example, SET1, SET7 and MLL methylate lysine 4 of histone H3, whereas Suv39h1, ESET and G9a specifically methylate lysine 9 of histone H3. Methylation at lysine 4 and lysine 9 are mutually exclusive and the epigenetic consequences of site-specific methylation are diametrically opposed: Methylation at lysine 4 correlates with an active state of transcription, whereas methylation at lysine 9 is associated with transcriptional repression and heterochromatin. Other lysine residues on histone H3 and histone H4 are also important sites of methylation by specific SET domain-containing enzymes. Although histones are the prime target of lysine methyltransferases, other cellular proteins carry N-methyllysine residues, including elongation factor 1A and the calcium sensing protein calmodulin. [9]

N-terminal methylation

Many eukaryotic proteins are post-translationally modified on their N-terminus. A common form of N-terminal modification is N-terminal methylation (Nt-methylation) by N-terminal methyltransferases (NTMTs). Proteins containing the consensus motif H2N-X-Pro-Lys- (where X can be Ala, Pro or Ser) after removal of the initiator methionine (iMet) can be subject to N-terminal α-amino-methylation. [10] Monomethylation may have slight effects on α-amino nitrogen nucleophilicity and basicity, whereas trimethylation (or dimethylation in case of proline) will result in abolition of nucleophilicity and a permanent positive charge on the N-terminal amino group. Although from a biochemical point of view demethylation of amines is possible, Nt-methylation is considered irreversible as no N-terminal demethylase has been described to date. [10] Histone variants CENP-A and CENP-B have been found to be Nt-methylated in vivo. [10]

Prenylcysteine

Eukaryotic proteins with C-termini that end in a CAAX motif are often subjected to a series of posttranslational modifications. The CAAX-tail processing takes place in three steps: First, a prenyl lipid anchor is attached to the cysteine through a thioester linkage. Then endoproteolysis occurs to remove the last three amino acids of the protein to expose the prenylcysteine α-COOH group. Finally, the exposed prenylcysteine group is methylated. The importance of this modification can be seen in targeted disruption of the methyltransferase for mouse CAAX proteins, where loss of isoprenylcysteine carboxyl methyltransferase resulted in mid-gestation lethality. [11]

The biological function of prenylcysteine methylation is to facilitate the targeting of CAAX proteins to membrane surfaces within cells. Prenylcysteine can be demethylated and this reverse reaction is catalyzed by isoprenylcysteine carboxyl methylesterases. CAAX box containing proteins that are prenylcysteine methylated include Ras, GTP-binding proteins, nuclear lamins and certain protein kinases. Many of these proteins participate in cell signaling, and they utilize prenylcysteine methylation to concentrate them on the cytosolic surface of the plasma membrane where they are functional. [11]

Methylations on the C-terminus can increase a protein's chemical repertoire [12] and are known to have a major effect on the functions of a protein. [1]

Protein phosphatase 2

In eukaryotic cells, phosphatases catalyze the removal of phosphate groups from tyrosine, serine and threonine phosphoproteins. The catalytic subunit of the major serine/threonine phosphatases, like Protein phosphatase 2 is covalently modified by the reversible methylation of its C-terminus to form a leucine carboxy methyl ester. Unlike CAAX motif methylation, no C-terminal processing is required to facilitate methylation. This C-terminal methylation event regulates the recruitment of regulatory proteins into complexes through the stimulation of protein–protein interactions, thus indirectly regulating the activity of the serine-threonine phosphatases complex. [13] Methylation is catalyzed by a unique protein phosphatase methyltransferase. The methyl group is removed by a specific protein phosphatase methylesterase. These two opposed enzymes make serine-threonine phosphatases methylation a dynamic process in response to stimuli. [13]

L-isoaspartyl

Damaged proteins accumulate isoaspartyl which causes protein instability, loss of biological activity and stimulation of autoimmune responses. The spontaneous age-dependent degradation of L-aspartyl residues results in the formation of a succinimidyl intermediate, a succinimide radical. This is spontaneously hydrolyzed either back to L-aspartyl or, in a more favorable reaction, to abnormal L-isoaspartyl. A methyltransferase dependent pathway exists for the conversion of L-isoaspartyl back to L-aspartyl. To prevent the accumulation of L-isoaspartyl, this residue is methylated by the protein L-isoaspartyl methyltransferase, which catalyzes the formation of a methyl ester, which in turn is converted back to a succinimidyl intermediate. [14] Loss and gain of function mutations have unmasked the biological importance of the L-isoaspartyl O-methyltransferase in age-related processes: Mice lacking the enzyme die young of fatal epilepsy, whereas flies engineered to over-express it have an increase in life span of over 30%. [14]

Physical effects

A common theme with methylated proteins, as with phosphorylated proteins, is the role this modification plays in the regulation of protein–protein interactions. The arginine methylation of proteins can either inhibit or promote protein–protein interactions depending on the type of methylation. The asymmetric dimethylation of arginine residues in close proximity to proline-rich motifs can inhibit the binding to SH3 domains. [15] The opposite effect is seen with interactions between the survival of motor neurons protein and the snRNP proteins SmD1, SmD3 and SmB/B', where binding is promoted by symmetric dimethylation of arginine residues in the snRNP proteins. [16]

A well-characterized example of a methylation dependent protein–protein interaction is related to the selective methylation of lysine 9, by SUV39H1 on the N-terminal tail of the histone H3. [9] Di- and tri-methylation of this lysine residue facilitates the binding of heterochromatin protein 1 (HP1). Because HP1 and Suv39h1 interact, it is thought the binding of HP1 to histone H3 is maintained and even allowed that to spread along the chromatin. The HP1 protein harbors a chromodomain which is responsible for the methyl-dependent interaction between it and lysine 9 of histone H3. It is likely that additional chromodomain-containing proteins will bind the same site as HP1, and to other lysine methylated positions on histones H3 and Histone H4. [13]

C-terminal protein methylation regulates the assembly of protein phosphatase. Methylation of the protein phosphatase 2A catalytic subunit enhances the binding of the regulatory B subunit and facilitates holoenzyme assembly. [13]

Related Research Articles

<span class="mw-page-title-main">Histone</span> Protein family around which DNA winds to form nucleosomes

In biology, histones are highly basic proteins abundant in lysine and arginine residues that are found in eukaryotic cell nuclei and in most Archaeal phyla. They act as spools around which DNA winds to create structural units called nucleosomes. Nucleosomes in turn are wrapped into 30-nanometer fibers that form tightly packed chromatin. Histones prevent DNA from becoming tangled and protect it from DNA damage. In addition, histones play important roles in gene regulation and DNA replication. Without histones, unwound DNA in chromosomes would be very long. For example, each human cell has about 1.8 meters of DNA if completely stretched out; however, when wound about histones, this length is reduced to about 90 micrometers (0.09 mm) of 30 nm diameter chromatin fibers.

Methylation, in the chemical sciences, is the addition of a methyl group on a substrate, or the substitution of an atom by a methyl group. Methylation is a form of alkylation, with a methyl group replacing a hydrogen atom. These terms are commonly used in chemistry, biochemistry, soil science, and biology.

<span class="mw-page-title-main">Histone methyltransferase</span> Histone-modifying enzymes

Histone methyltransferases (HMT) are histone-modifying enzymes, that catalyze the transfer of one, two, or three methyl groups to lysine and arginine residues of histone proteins. The attachment of methyl groups occurs predominantly at specific lysine or arginine residues on histones H3 and H4. Two major types of histone methyltranferases exist, lysine-specific and arginine-specific. In both types of histone methyltransferases, S-Adenosyl methionine (SAM) serves as a cofactor and methyl donor group.
The genomic DNA of eukaryotes associates with histones to form chromatin. The level of chromatin compaction depends heavily on histone methylation and other post-translational modifications of histones. Histone methylation is a principal epigenetic modification of chromatin that determines gene expression, genomic stability, stem cell maturation, cell lineage development, genetic imprinting, DNA methylation, and cell mitosis.

<span class="mw-page-title-main">Histone H3</span> One of the five main histone proteins

Histone H3 is one of the five main histones involved in the structure of chromatin in eukaryotic cells. Featuring a main globular domain and a long N-terminal tail, H3 is involved with the structure of the nucleosomes of the 'beads on a string' structure. Histone proteins are highly post-translationally modified however Histone H3 is the most extensively modified of the five histones. The term "Histone H3" alone is purposely ambiguous in that it does not distinguish between sequence variants or modification state. Histone H3 is an important protein in the emerging field of epigenetics, where its sequence variants and variable modification states are thought to play a role in the dynamic and long term regulation of genes.

<span class="mw-page-title-main">Histone H4</span> One of the five main histone proteins involved in the structure of chromatin

Histone H4 is one of the five main histone proteins involved in the structure of chromatin in eukaryotic cells. Featuring a main globular domain and a long N-terminal tail, H4 is involved with the structure of the nucleosome of the 'beads on a string' organization. Histone proteins are highly post-translationally modified. Covalently bonded modifications include acetylation and methylation of the N-terminal tails. These modifications may alter expression of genes located on DNA associated with its parent histone octamer. Histone H4 is an important protein in the structure and function of chromatin, where its sequence variants and variable modification states are thought to play a role in the dynamic and long term regulation of genes.

Histone methylation is a process by which methyl groups are transferred to amino acids of histone proteins that make up nucleosomes, which the DNA double helix wraps around to form chromosomes. Methylation of histones can either increase or decrease transcription of genes, depending on which amino acids in the histones are methylated, and how many methyl groups are attached. Methylation events that weaken chemical attractions between histone tails and DNA increase transcription because they enable the DNA to uncoil from nucleosomes so that transcription factor proteins and RNA polymerase can access the DNA. This process is critical for the regulation of gene expression that allows different cells to express different genes.

<span class="mw-page-title-main">Methyltransferase</span> Group of methylating enzymes

Methyltransferases are a large group of enzymes that all methylate their substrates but can be split into several subclasses based on their structural features. The most common class of methyltransferases is class I, all of which contain a Rossmann fold for binding S-Adenosyl methionine (SAM). Class II methyltransferases contain a SET domain, which are exemplified by SET domain histone methyltransferases, and class III methyltransferases, which are membrane associated. Methyltransferases can also be grouped as different types utilizing different substrates in methyl transfer reactions. These types include protein methyltransferases, DNA/RNA methyltransferases, natural product methyltransferases, and non-SAM dependent methyltransferases. SAM is the classical methyl donor for methyltransferases, however, examples of other methyl donors are seen in nature. The general mechanism for methyl transfer is a SN2-like nucleophilic attack where the methionine sulfur serves as the leaving group and the methyl group attached to it acts as the electrophile that transfers the methyl group to the enzyme substrate. SAM is converted to S-Adenosyl homocysteine (SAH) during this process. The breaking of the SAM-methyl bond and the formation of the substrate-methyl bond happen nearly simultaneously. These enzymatic reactions are found in many pathways and are implicated in genetic diseases, cancer, and metabolic diseases. Another type of methyl transfer is the radical S-Adenosyl methionine (SAM) which is the methylation of unactivated carbon atoms in primary metabolites, proteins, lipids, and RNA.

The family of heterochromatin protein 1 (HP1) consists of highly conserved proteins, which have important functions in the cell nucleus. These functions include gene repression by heterochromatin formation, transcriptional activation, regulation of binding of cohesion complexes to centromeres, sequestration of genes to the nuclear periphery, transcriptional arrest, maintenance of heterochromatin integrity, gene repression at the single nucleosome level, gene repression by heterochromatization of euchromatin, and DNA repair. HP1 proteins are fundamental units of heterochromatin packaging that are enriched at the centromeres and telomeres of nearly all eukaryotic chromosomes with the notable exception of budding yeast, in which a yeast-specific silencing complex of SIR proteins serve a similar function. Members of the HP1 family are characterized by an N-terminal chromodomain and a C-terminal chromoshadow domain, separated by a hinge region. HP1 is also found at some euchromatic sites, where its binding can correlate with either gene repression or gene activation. HP1 was originally discovered by Tharappel C James and Sarah Elgin in 1986 as a factor in the phenomenon known as position effect variegation in Drosophila melanogaster.

<span class="mw-page-title-main">Methyllysine</span> Derivative of the amino acid residue lysine

Methyllysine is derivative of the amino acid residue lysine where the sidechain ammonium group has been methylated one or more times.

<span class="mw-page-title-main">Histone-modifying enzymes</span> Type of enzymes

Histone-modifying enzymes are enzymes involved in the modification of histone substrates after protein translation and affect cellular processes including gene expression. To safely store the eukaryotic genome, DNA is wrapped around four core histone proteins, which then join to form nucleosomes. These nucleosomes further fold together into highly condensed chromatin, which renders the organism's genetic material far less accessible to the factors required for gene transcription, DNA replication, recombination and repair. Subsequently, eukaryotic organisms have developed intricate mechanisms to overcome this repressive barrier imposed by the chromatin through histone modification, a type of post-translational modification which typically involves covalently attaching certain groups to histone residues. Once added to the histone, these groups elicit either a loose and open histone conformation, euchromatin, or a tight and closed histone conformation, heterochromatin. Euchromatin marks active transcription and gene expression, as the light packing of histones in this way allows entry for proteins involved in the transcription process. As such, the tightly packed heterochromatin marks the absence of current gene expression.

<span class="mw-page-title-main">CBX5 (gene)</span> Protein-coding gene in humans

Chromobox protein homolog 5 is a protein that in humans is encoded by the CBX5 gene. It is a highly conserved, non-histone protein part of the heterochromatin family. The protein itself is more commonly called HP1α. Heterochromatin protein-1 (HP1) has an N-terminal domain that acts on methylated lysines residues leading to epigenetic repression. The C-terminal of this protein has a chromo shadow-domain (CSD) that is responsible for homodimerizing, as well as interacting with a variety of chromatin-associated, non-histone proteins.

<span class="mw-page-title-main">Tudor domain</span>

In molecular biology, a Tudor domain is a conserved protein structural domain originally identified in the Tudor protein encoded in Drosophila. The Tudor gene was found in a Drosophila screen for maternal factors that regulate embryonic development or fertility. Mutations here are lethal for offspring, inspiring the name Tudor, as a reference to the Tudor King Henry VIII and the several miscarriages experienced by his wives.

<span class="mw-page-title-main">SET domain</span>

The SET domain is a protein domain that typically has methyltransferase activity. It was originally identified as part of a larger conserved region present in the Drosophila Trithorax protein and was subsequently identified in the Drosophila Su(var)3-9 and 'Enhancer of zeste' proteins, from which the acronym SET is derived [Su(var)3-9, Enhancer-of-zeste and Trithorax].

H3K4me3 is an epigenetic modification to the DNA packaging protein Histone H3 that indicates tri-methylation at the 4th lysine residue of the histone H3 protein and is often involved in the regulation of gene expression. The name denotes the addition of three methyl groups (trimethylation) to the lysine 4 on the histone H3 protein.

H3K9me3 is an epigenetic modification to the DNA packaging protein Histone H3. It is a mark that indicates the tri-methylation at the 9th lysine residue of the histone H3 protein and is often associated with heterochromatin.

H3K36me3 is an epigenetic modification to the DNA packaging protein Histone H3. It is a mark that indicates the tri-methylation at the 36th lysine residue of the histone H3 protein and often associated with gene bodies.

H3K14ac is an epigenetic modification to the DNA packaging protein Histone H3. It is a mark that indicates the acetylation at the 14th lysine residue of the histone H3 protein.

H3K36me is an epigenetic modification to the DNA packaging protein Histone H3, specifically, the mono-methylation at the 36th lysine residue of the histone H3 protein.

H3R42me is an epigenetic modification to the DNA packaging protein histone H3. It is a mark that indicates the mono-methylation at the 42nd arginine residue of the histone H3 protein. In epigenetics, arginine methylation of histones H3 and H4 is associated with a more accessible chromatin structure and thus higher levels of transcription. The existence of arginine demethylases that could reverse arginine methylation is controversial.

H3R8me2 is an epigenetic modification to the DNA packaging protein histone H3. It is a mark that indicates the di-methylation at the 8th arginine residue of the histone H3 protein. In epigenetics, arginine methylation of histones H3 and H4 is associated with a more accessible chromatin structure and thus higher levels of transcription. The existence of arginine demethylases that could reverse arginine methylation is controversial.

References

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