Posttranslational modification is the chemical modification of a protein after its translationchemical proteintranslation

Proteins often undergo several post-translational modification steps in parallel to protein folding. These modifications can be transient or permanent. Most modifications are susceptible to alteration during the lifespan of proteins. Post-translational modifications generate variability in proteins that are far beyond that provided by the genetic code. Co- and post-translational modifications can convert the 20 specific codon-encoded amino acids into more than 100 variant amino acids with new properties. These, and a number of other modifications, can considerably increase the information content and functional repertoire of proteins

Post-Translational Modification 1. Dealing with the N-terminal residue In bacteria, the N-terminal residue of the newly- synthesized protein is modified to remove the formyl group. The N-terminal methionine may also be removed. In some cases the carboxy terminal residues are removed enzymatically 2. Loss of signal sequences residues at the amino terminal end of some proteins play a role in directing the protein to its ultimate destination in the cell. Signal sequences are ultimately removed by specific peptidases

Post-Translational Modification 3. Amino Acid Modifications Many of the amino-acid side-chains can be modified Acetylation The amino-terminal residues of some proteins are acetylated. This is more common in eukaryotes than prokaryotes e.g. the N-terminal serine of histone H4 is invariably acetylated as are a number of lysine residues.

Phosphorylation In prokaryotes phosphorylation has been well established and shown to play an essential role in the control of bacterial protein function esp. enzyme activation. E.g. the activity of isocitrate dehydrogenase is regulated via its reversible phosphorylation, which modulates the partition of carbon flux between the Krebs cycle and the glyoxylate bypass Phosphorylation of proteins (at Ser, Thr, Tyr and His residues) is an important regulatory mechanism. For example, the activity of glycogen phosphorylase is regulated by phosphorylation of Serine 14. Phosphorylation of tyrosine residues is an important aspect of signal transduction pathways. Bacterial cells sense and respond to environmental signals through histidine phosphorylation

Methylation In bacterial chemotaxis where in the absence of an added stimulus chemotaxis proteins are methylated to a basal level Methylation of the 50S ribosomal proteins from Bacillus stearothermophilus, Bacillus subtilis, Alteromonas espejiana, and Halobacterium cutirubrum The activity of histones can be modified by methylation. Lysine 20 of histone H4 can be mono- or di- methylated.

Carboxylation Not very important in prokaryotes The blood coagulation factor, prothrombin, contains a large number of carboxylated glutamatic acid residues in the N-terminal 32 amino acids. These modified residues are essential for activity. The modification requires vitamin K.

Hydroxylation This does not occur in prokaryotes and can lead to problems with the expression of recombinant proteins The conversion of proline to hydroxyproline in collagen is the classical example of a post-translational modification

Glycosylation Numerous virulence factors of bacterial pathogens have been found to be covalently modified with carbohydrate residues, thereby identifying these factors as true glycoproteins. In several bacterial species, gene clusters suggested to represent a general protein glycosylation system have been identified. In other cases, genes encoding highly specific glycosyltransferases have been found to be directly linked with virulence genes. Seems may be a role for glycosylation in pathogenesis. both O-linked and N-linked protein glycosylation pathways in bacteria, particularly amongst mucosal-associated pathogens Many extracellular (but not intracellular) proteins are glycosylated. Mono- or Oligo-saccharides can be attached to asparagine (N- linked) or to serine/threonine (O-linked) residues.

Nucleotidylation Mononucleotide addition is used to regulate the activity of some enzymes. Two different examples are found among the system that regulates Nitrogen utilization in E. coli: Glutamine synthetase is adenylylated (i.e. AMP is added) at a specific tyrosine residue. The enzyme is inactive when it is adenylylated. The degree of adenylylation is controlled by a regulatory protein, PII. The ability of PII to regulate the adenylylation of glutamine synthetase is in turn regulated by its own uridylylation (i.e. the covalent addition of UMP). PII is also uridylylated at a tyrosine residue.

4. Lipid Addition Some proteins have lipid moieties attached: The viral src protein is myristoylated at the N-terminal glycine. Rhodoposin is palmitoylated at a cysteine residue The ras oncogene protein is farnesylated as are some G proteins. Some eukaryotes, notably parasitic protozoa, have glycosylphosphatidylinositol-linked proteins. 5. Adding Prosthetic Groups Proteins that require a prosthetic group for activity must have this group added. For example, the haem (heme) group must be added to globins and cytochromes; Fe-S clusters must be added to ferredoxins. 6. Forming Disulfide Bonds Many extracellular proteins contain disulfide cross-links (intracellular proteins almost never do). The cross-links can only be established after the protein has folded up into the correct shape. The formation of disulfide bonds is aided by the enzyme protein disulfide isomerase in eukaryotes and by the DsbA protein in bacteria.

7. Proteolytic Processing Some proteins are synthesized as inactive precursor polypeptides which become activated only after proteolytic cleavage of the precursor polypeptide chain. E.g. Chymotrypsin & Trypsin These are both synthesized as zymogens. Cleavage of chymotrypsinogen between Arg15 and Ile 16 by trypsin yields the enzymatically active pi-chymotrypsin. Two further proteolytic cleavages catalyzed by chymotrypsin removes the dipeptides Ser14- Arg15 and Thr147-Asn148 to yield alpha- chymotrypsin.