Polypeptides frequently undergo a variety of enzymatic chemical modifications, during or after translation. The modifications involve covalent attachment of simple or complex chemical groups, and they may be reversible (as a way of changing the behavior of the protein toward different outcomes) or be irreversible. The dbPTM database provides a searchable database of protein translation modifications.
Many proteins have one or more simple chemical groups attached to specific amino acids. In addition, more complex chemical groups are covalently attached to the poly peptide backbone of certain proteins. In the latter case, large groups may be added irreversibly: carbohydrates to secreted proteins, and lipids and glycolipids to many membrane proteins. In other cases, ADP-ribose units and certain types of proteins can be reversibly attached to proteins to regulate them in some way (Table 1).
Table1. MAJOR TYPES OF CHEMICAL MODIFICATION IN PROTEINS
Another type of post-translational processing involves specific cleavage of a precursor polypeptide to yield one or more active polypeptide products.
Addition of simple chemical groups
The characteristics and functions of proteins are often regulated by reversible attachment of very simple chemical groups, such as phosphate, methyl, acetyl, hydroxyl, and carboxyl groups. Small armies of enzymes are needed to add or remove these groups from specific proteins, or protein classes, such as designated kinases, methylases, and acetylases to add phosphate, methyl, and acetyl groups, respectively, and phosphatases, demethylases, and deacetylases to remove them.
These modifications are essential for diverse cell functions including regulation of chromatin structure, transcription, cell signaling, and so on, as described in later chapters, and key proteins are known to undergo a wide range of post-translational modifications (Figure 1).
Fig1. Post-translational modifications in the p53 protein that are known to be responsible for specific changes in its behaviour. The p53 protein is known to undergo many different post-translational modifications, but only the ones known to be directly responsible for the biological effects listed to the left are shown here. GlcNAc, N-acetylglucosamine. (Adapted from Gu B & Zhu WG [2012] Int J Biol Sci 8:672–684; PMID 22606048. With permission from Ivyspring International Publisher.)
Addition of carbohydrate groups
Glycoproteins have oligosaccharides covalently attached to the side chains of certain amino acids. Few proteins in the cytosol are glycosylated (carry an attached carbohydrate); if they are, they have a single sugar residue, N-acetylglucosamine, attached to a serine or threonine residue. However, proteins that are secreted from cells or trans ported to lysosomes, the Golgi apparatus, or the plasma membrane are routinely glycosylated. In these cases, the sugars are assembled as oligosaccharides before being attached to the protein.
Two major types of glycosylation are recognized. Carbohydrate N-glycosylation involves attaching a carbohydrate group to the nitrogen atom of an asparagine side chain and O-glycosylation entails adding a carbohydrate to the oxygen atom of an OH group carried by the side chains of certain amino acids (see Table 1). Some modifications simply involve adding monosaccharides or disaccharides, but in others more complex carbohydrates are attached that may be linear or branched polymers.
Proteoglycans are proteins with attached glycosaminoglycans (polysaccharides) that usually include repeating disaccharide units containing glucosamine or galactosamine. The best-characterized proteoglycans are components of the extracellular matrix, a complex network of macromolecules secreted by, and surrounding, cells in tissues or in culture systems.
Addition of lipid and glycolipid groups
Some proteins, notably membrane proteins, are modified by the addition of fatty acyl or prenyl groups. The added groups typically serve as membrane anchors, hydrophobic amino acid sequences that secure a newly synthesized protein within either a plasma membrane or the endoplasmic reticulum (see Table 1).
Anchoring of a protein to the outer layer of the plasma membrane often involves attaching a glycosylphosphatidylinositol (GPI) group (Figure 2A). This glycolipid group contains fatty acyl groups that serve as the membrane anchor; they are linked successively to a glycerophosphate unit, an oligosaccharide unit, and finally through a phosphoethanolamine unit to the C-terminus of the protein. The entire protein, except for the GPI anchor, is located in the extracellular space.
Fig2. Examples of protein modification by attachment of complex groups. (A) Addition of glycosylphosphatidylinositol (GPI) to anchor a protein to the surface of the plasma membrane. The C-terminal carboxyl group of the protein is attached by an ethanolamine group (green) to a glycan group (blue), based on mannose (MAN) and glucose (GLU), that is linked in turn to a phosphatidylinositol group (red). The latter has long acyl side groups that insert into the plasma membrane to provide anchorage. (B) Addition of ADP-ribose units. The figure shows the chemical structures of nicotinamide, nicotinamide adenine dinucleotide (NAD+), and poly(ADP-ribose). Addition of ADP-ribose units is achieved using NAD+ as a donor of ADP-ribose. Poly(ADP-ribose) is a branched polymer synthesized on acceptor proteins by poly(ADP-ribose) polymerases (PARPs) using NAD+ as a donor of ADP-ribose units. (A, modified from Rosse WF & Ware RE [1995] Blood 86:3277–3286; PMID 7579428; B, adapted from Luo X & Kraus WL [2012] Genes Dev 26:417–432; PMID 22391446. With permission from Cold Spring Harbor Laboratory Press.)
Addition of proteins
Some proteins are regulated by attachment of specialized proteins. Sumoylation is a protein modification in which SUMO (small ubiquitin-like modifier) proteins are reversibly attached to proteins. Sumoylation induces the proteins to change their behavior (via altered binding properties, change of localization within cells, and so on), and is important in regulating certain processes, such as transcription, chromatin structure, DNA repair, protein stability, transport between the nucleus and cytoplasm, and so on.
Ubiquitin proteins resemble SUMO proteins and can also be reversibly attached to proteins. A major purpose of this modification is to target proteins for destruction (when the protein needs to be replaced, or is a threat to the cell). Adding a small chain of ubiquitin residues (polyubiquitin) to a protein marks that protein for proteolytic degradation in the proteasome, whereupon the polyubiquitin is recycled as single ubiquitin units. In other cases, a single ubiquitin residue can be attached to a protein and this type of ubiquitin modification can have different regulatory roles.
ADP-ribosylation
Another reversible protein modification involves the enzymatic addition of ADP-ribose units, which are donated by nicotinamide adenine dinucleotide (NAD+). Mono-ADP-ribosyltransferases catalyze the addition of a single ADP-ribose unit from NAD+ to an arginine side chain of the target protein. Poly(ADP-ribose) polymerases (PARPs) catalyze the addition of multiple ADP-ribose units provided by donor NAD+ molecules (Figure 2B).
Post-translational cleavage
The primary translation product may also undergo internal cleavage to generate a smaller mature product. Occasionally the initiating methionine is cleaved from the primary translation product, as during the synthesis of β-globin. Secreted proteins (plasma proteins, polypeptide hormones, neuropeptides, growth factors, and so on) are typically produced with a short N-terminal signal sequence that serves as a destination tag and is cleaved prior to export from the cell. Post-translational cleavage is sometimes also employed to generate active polypeptides from a larger polypeptide precursor (see the example of insulin production in Figure 3).
Fig3. Insulin synthesis involves multiple post-translational cleavages of polypeptide precursors. Human insulin mRNA is translated to give a 110-amino acid (aa) preproinsulin that has a 24-aa N-terminal leader sequence, an address tag required for the protein to be exported from the cell. In processing, the leader sequence is cleaved off and discarded. The remaining 86-aa proinsulin precursor contains a central segment (the connecting peptide) that may maintain the conformation of the A and B chains of insulin in readiness for making the final insulin protein. At the last moment, the connecting peptide is excised and the A and B chains are then covalently bonded together by disulfide bridges.
