Posttranslational covalent modifications

The above definition of molecular chaperone is entirely fnnctional and contains no constraints on the mechanisms by which different chaperones may act. The term noncovalent is nsed to exclude those proteins that carry out posttranslational covalent modifications. Protein disulfide isomerise may seem to be an exception, bnt it is both a covalent modification enzyme and a molecular chaperone. It is helpful to think of a molecnlar chaperone as a fnnction rather than as a molecnle. Thns, no reason exists why a chaperone function shonld not be a property of the same molecnle that has other fnnctions. Other examples include peptidyl-prolyl isomerase, which possesses both enzymatic and chaperone activities in different regions of the molecnle, and the alpha-crystallins, which combine two essential fnnctions in the same molecnle in the lens of the eye-contribnting to the transparency and the refractive index reqnired for vision as well... 

The step labeled p in Figure 1 represents modification of primary proteins to render them functional examples would be posttranslational covalent modifications (e.g., phosphorylation) and binding with other proteins or other molecules. Represented within the set of steps p are the many regulatory events (other than transcription and translation) affecting gene expression and the overall physiology of the cell. 

R. Uy and F. Wold, Posttranslation covalent modification of proteins. Science 198,890 (1977). 

Figure 1 Covalent modifications of DNA and histones play a fundamental role in the regulation of differentiation and development. The writers, readers, and erasers of this dynamic code are potentially amenable to modulation with small molecules. Lysine methylation is a critical posttranslational modification influencing chromatin function (PMT = protein lysine methyltransferase, royal family proteins bind KMe, KDM = lysine demethylase).

The third category of posttranslational reactions, those involved in covalent modification of the amino acid side chains, is by far the largest. According to the data in Fig. 1 (and Ref. 2) there are about 98 known derivatives of amino acid side chains in proteins. In the following paragraphs some of these will be discussed briefly. 

Posttranslational modifications are enzyme-catalyzed covalent modifications of a mature protein after it has been synthesized. Examples of posttranslational modifications are phosphorylation, glycosylation, sulfation, methylation and prenylation. Espedally those modifications that are reversible, such as phosphorylation by de-phosphorylation through the action of phosphatases are important in regulation. 

Posttranslational Covalent Protein Modification by NO-derived Species 9... 

Posttranslational Protein Modification a mechanism of biological control of protein activity by covalent modification after protein synthesis... 

POSTTRANSLATIONAL COVALENT PROTEIN MODIFICATION BY. NO-DERIVED SPECIES... 

The dioxygen reduction site of the key respiratory enzyme, cytochrome c oxidase [E.C. 1.9.3.1], is a bimetallic catalytic center comprised of a heme iron adjacent to a Type 2 mononuclear copper center (see Cytochrome Oxidase). The recent solution of the X-ray crystal structure of this enzyme revealed an entirely unanticipated covalent modification of the protein structure, a cross-link between a histidine and tyrosine side chain (23) within the active site (Figure 2)." This extraordinary posttranslational modification has been confirmed by peptide mapping and mass spectrometry, and has been detected as a conserved element in cytochrome c oxidases isolated from organisms ranging from bacteria to cows. The role of the cross-linked structure in the function of cytochrome c oxidase is still controversial." " ... 

N is often limiting in the marine environment. Further, many enzymes are sensitive to cellular substrate concentrations rather than extracellular concentrations and it is difficult to measure the relevant intracellular metabohte pools. In vitro assays may affect the conformation of enzymes and the degree to which they are modified. For example, allosteric effects (see Section 1.3.3) may be modified under in vitro conditions. Many enzymes undergo posttranslational regulation wherein enzyme activity is affected by binding of activator/inactivator proteins and covalent modification of the enzyme (e.g., adenylylation, phosphorylation or carbamylation) (Ottaway, 1988). When there is posttranslational modification of enzymes, enzyme activity measured in assays may be unrelated to in vivo activity (see Section 2.2.1) and there are few ways to determine the extent of enzyme modification in nature. 

Another approach to regulating the activity of transcription factors is to modify the chromatin state and therefore the accessibility of the promoter and TF binding sites (78, 79). Chromatin states are modulated partially by posttranslational modifications of the histones in nucleosomes, which have significant effects on transcription levels (14, 80). These modifications include phosphorylation, methylation, acetylation, and ubiquitination (80, 81). An early indication of the importance of these covalent modifications on transcription emerged from the observation... 

Polypeptide chains are synthesized in the cytoplasm of a cell by the process known as translation. The polypeptide may be ready for use immediately after translation or it may require further maturation steps, such as facilitated folding, complex formation, transport to another cell compartment, or covalent modification of chemical groups on the protein. The term posttranslational modifications is used for the last of these processes. Some of the modifications take place in the cytoplasm or nucleus, particularly phosphorylation and acetylation, and others take place in the endoplasmic reticulum or Golgi apparatus, particularly the addition of sugar or polysaccharide residues. 

Although lantibiotics are not enzymes and do not contain cofactors they are included here and briefly discussed because they are proteins whose biosynthesis requires extensive and unusual posttranslational modifications. These modifications endow the gene product with a new function, in this case an antimicrobial activity, which would not exist in the absence of these covalent modifications to the protein. They are found in peptides of microbial origin. Their name derives from lanthionine, which is a nonproteinogenic amino acid composed of two alanine residues that are crosslinked on their /3-carbon atoms by a thioether linkage (Figure 15). The wide array of fascinating posttranslational modifications which are present in this class of peptide antibiotics is reminiscent of those seen in some protein-derived cofactors. 

Hundreds of modifications to the protein backbone have been reported and reviewed." ° For many biopharmaceutical proteins, however, only a handful of modifications are usually detected. These include both posttranslational modifications that are results of intracellular enzymatic processes, and covalent modifications that occur during or after the manufacturing process, either induced by process conditions or resulting from degradation (discussed in Section II.A.4). 

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