Post-translational modifications, 62, Table

Technical advances facilitating genetic manipulation of animal cells now allow routine production of therapeutic proteins in such systems. The major advantage of these systems is their ability to carry out post-translational modification of the protein product. As a result, many biopharmaceuticals that are naturally glycosylated are now produced in animal cell lines (Table 3.9). Chinese hamster ovary (CHO) and baby hamster kidney (BHK) cells have become particularly popular in this regard. 

EMBL Nucleotide Sequence Database. SWISS-PROT consists of core sequence data with minimal redundancy, citation and extensive annotations including protein function, post-translational modifications, domain sites, protein structural information, diseases associated with protein deficiencies and variants. SWISS-PROT and TrEMBL are available at EBI site, http //www.ebi.ac.uk/swissprot/, and ExPASy site, http //www.expasy.ch/sprot/. From the SWISS-PROT and TrEMBL page of ExPASy site, click Full text search (under Access to SWISS-PROT and TrEMBL) to open the search page (Figure 11.3). Enter the keyword string (use Boolean expression if required), check SWISS-PROT box, and click the Submit button. Select the desired entry from the returned list to view the annotated sequence data in Swiss-Prot format. An output in the fasta format can be requested. Links to BLAST, feature table, some ExPASy proteomic tools (e.g., Compute pI/Mw, ProtParam, ProfileScan, ProtScale, PeptideMass, ScanProsite), and structure (SWISS-MODEL) are provided on the page. 

Table 1.1 Some Examples of Post-Translational Modification ...

Table I. In Vivo Nonhydrolytic Enzyme-Catalyzed Post-Translational Modifications of Amino Acid Residues of Proteins0...

Table 8.1 Post-translational modifications and corresponding average mass variations.

Proteins and peptides are linear polymers made up of combinations of the 20 most common amino acids linked with each other by peptide bonds. Moreover, the protein produced by the ribosome may undergo covalent modifications, called post-translational modifications, after its incorporation of amino acids. Over 200 such modifications have been detected already [13,14], the most important being glycosylation, the formation of disulfide bridges, phosphorylation, sulfation, hydroxylation, carboxylation and acetylation of the N-terminal acid [15]. The most frequent are listed in Table 8.1 and a more comprehensive database of mass changes due to post-translational modifications of peptides and proteins is available on the Internet [16]. 

Amino acid moieties of many proteins are enzymatically modified within the intracellular compartments following their incorporation into the polypeptide chains. For example, many asparagine residues in proteins of enkaryotic cells have carbohydrates attached to them, converting the protein into a glycoprotein. Several other post-translationa 1 modifications involving many of the 20 amino acids in proteins have been observed. Table 2.4 is a partial list of post-translational modifications found in mammalian proteins. 

Table 2.4. Common post-translational modifications in mammalian proteins...

Five transformants produced a major secreted protein band, with an estimated molecular weight of 40 kDa and no protein was detected in the control. The estimated molecular weight is greater than that predicted or observed from E. coli, which is likely due to post-translational modification. P6 and P10 transformants were retained to perform enzymatic assays. The culture supernatants were assays for activity against methyl caffeate (MCA) and methyl ferulate (MFA). The recombinant proteins were shown to be active as a feruloyl esterase and show the characteristics of a type B ferulic acid esterase.6 Feruloyl esterase activity is reported in Table 1. 

Besides AFl and AF2, 16 other Ascaris FMRFamide-like peptides have been isolated and sequenced (12 sequences are shown in Table 14.2) (194-197 Cowden and Stretton, unpublished). These sequences are unique and not related by post-translational modification or, in most cases, by proteolysis they are also different from FMRFamide-like sequences reported in other organisms, whether obtained from isolated and sequenced peptides, or by deduction from the DNA sequences of genes. It is clear that there is a family of FMRFamide-like peptides in A. suum. They can be divided into several subfamilies related by sequence. Initial experiments on their bioactivity suggest that these may also be functional subfamilies. Measurements of effects on muscle tension, and on input resistance of four types of motoneurons, show that there are at least four classes of biological activity controlled by these peptides AFl and AF2 form one class that causes contraction and the generation of rhythmic activity in muscle (194,197), AF3 and AF4 cause muscle contraction (195), AF5 and AF7 reduce the input resistance of both excitatory and inhibitory motoneurons, and AFl 1 relaxes muscle and increases the input resistance of inhibitory motoneurons (Davis and Stretton, unpublished). 

The cause of the difference in pi between CyPAg o and CyPAg gg is unknown, but it may be due to other types of post-translational modification because the A-terminal residue of CyPAg gg is free (Table II). 

A large number of proteins undergo post-translational modifications (Wold, 1981 Harding and Crabbe, 1992). Modifications of proteins post-translationally provide the means for targeting/translocation of many proteins and regnlation of protein activities/functions. Table 13.10 summarizes common post-translational modifications that regulate enzyme activities. 

TABLE 16.9 Online resources for functional and post-translational modification sites... 

Table 2. Some amino acid derivatives formed by post-translational modification in polypeptides. For references see H.B. Vickery [11]...

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