Collagen posttranslational modifications

Many other peptides are synthesized as proproteins that require modifications before attaining biologic activity. Many of the posttranslational modifications involve the removal of amino terminal amino acid residues by specific aminopeptidases. Collagen, an abundant protein in the extracellular spaces of higher eukaryotes, is synthesized as procollagen. Three procol-... 

Prolyl 4-hydroxylation is the most abundant posttranslational modification of collagens. 4-Hydroxylation of proline residues increases the stability of the triple helix and is a key element in the folding of the collagen triple helix. " In vertebrates, almost all the Yaa position prolines of the Gly-Xaa-Yaa repeat are modified to 4(I( )-hydroxylproline by the enzyme P4H (EC 1.14.11.2), a member of Fe(II)- and 2-oxoglutarate-dependent dioxygenases. This enzyme is an 0 2/ b2-type heterotetramer in which the / subunit is PDI (EC 5.3.4.1), which is a ubiquitous disulfide bond catalyst. The P4H a subunit needs the 13 subunit for solubility however, the 13 subunit, PDI, is soluble by itself and is present in excess in the ER. Three isoforms of the a subunit have been identified and shown to combine with PDI to form [a(I)]2/ 2) [< (II)]2/32> or [a(III)]2/32 tetramers, called the type... 

Hydroxyproline and hydroxylysine Collagen contains hydroxy proline (hyp) and hydroxylysine (hyl), which are not present in most other proteins. These residues result from the hydroxylation of some of the proline and lysine residues after their incorporation into polypeptide chains (Figure 4.6). The hydroxylation is, thus, an example of posttranslational modification (see p. 440). Hydroxy proline is important in stabilizing the triple-helical structure of colla gen because it maximizes interchain hydrogen bond formation. 

Posttranslational modification Examples of posttranslational modification POSTTRANSLATIONAL MODIFICATION OF POLYPEPTIDE CHAINS (p. 440) Many polypeptide chains are covalently modified after translation. Such modifications include trimming excess amino acids, phosphorylation which may activate or inactivate the protein, glycosylation which targets a protein to become part of a plasma membrane or lysosome or be secreted from the cell, or hydroxylation such as that seen in collagen. 

Hydroxyproline is formed by a posttranslational modification of proline residues in the protein. The 14C-labeled hydroxyproline is not incorporated directly into the collagen because there is no genetic codon to specify the incorporation of hydroxyproline. 

A second posttranslation modification that occurs in the maturation of collagen is the oxygenation of lysine residues to form 5-e/yfAro-hydroxylysine, which serves as an anchor for attachment of carbohydrates in a glycosylation process (equation 2). 

Biologic Background Structure of the Collagen Triple Helix Posttranslational Modifications Stability of the Collagen Triple Helix... 

Figure 3 Posttranslational modifications in collagen, (a) 3(S)-hydroxyproline. (b) 4(R)-hydroxyproline. (c) 5-hydroxylysine. (d) O-p-galactosyl-5-hydroxylysine. (e) 2-0-a-D-glucosyl-0-p-D-galactosyl-5-hydroxylysine.

We have identified a known posttranslational modification, hydroxylysine, in unexpected places. This modification is only partial, typically 5-25%, at one site only in tPA (at Lys /), rCD4 (at Lys ) and rCD4-IgG (at Lys ). All of the observed Hyl sites were foxmd in Xaa-Hyl-Gly sequences, as is the case for the known Hyl sites in collagens. The modification appears to be cell line-independent, as it was found in tPA from Bowes melanoma cells as well as rtPA from transfected CHO and 293 cells. 

FIGURE 7.1 Posttranslational modifications in collagen. (From Sikorski, Z.E., Proteins, in Chemical and Functional Properties of Food Components, Sikorski, Z.E., Ed., Technomic Publishing Co. Inc., PA, 1997. With permission.)... 

Synthesis of structurally altered collagen due to defects at the level of DNA transcription, RNA processing, translation, or posttranslational modifications. 

The formation of interchain disulfide linkages in the C-terminal propeptide. The latter cannot be formed until translation is nearly finished. The rate of disulfide bond and triple-helix formation varies greatly from one cell type to another—only minutes in tendon cells that synthesize type I collagen but an hour in cells that synthesize basement membrane collagen. These differences in synthesis time may account for the variations in hydroxylation and glycosylation. Thus, the extent of posttranslational modifications depends not only on levels of enzyme and cofactors but also on the time available. 

Pyridinium cross-links (pyridinoline and deoxypyridinoline) Posttranslational modification of lysine and hydroxylysine residues of collagen... 

Lysine tyrosylquinone (LTQ) (Figure 3) is the protein-derived cofactor of mammalian lysyl oxidase, an important enzyme in the metabolism of connective tissue. Lysyl oxidase catalyzes the posttranslational modification of elastin and collagen. It oxidizes selected peptidyl lysine residues to peptidyl a-aminoadipic -semialdehyde residues. This initiates formation of the covalent cross-linkages that insolubilize these extracellular proteins. This enzyme also contains copper as a second prosthetic group. 

Hydroxyproline residues are generated by posttranslational modification, following completion of the polypeptide chain. The nonhydroxylated collagen precursor is called procollagen (Figure 6.14, Figure... 

Collagen, a family of fibrous proteins, is produced by a variety of cell types but principally by fibroblasts (cells found in interstitial connective tissue), muscle cells, and epithelial cells. Type I collagen [collagen(l)], the most abundant protein in mammals, is a fibrous protein that is the major component of connective tissue. It is found in the extracellular matrix (ECM) of loose connective tissue, bone, tendons, skin, blood vessels, and the cornea of the eye. Collagen(l) contains approximately 33% glycine and 21% proline and hydroxyproline. Hydroxyproline is an amino acid produced by posttranslational modification of peptidyl proline residues (see Chapter 7, section V.C., for an earlier introduction to collagen). 

Many other amino acids, in addition to the ones listed here, are known to exist. They occur in some, but by no means all, proteins. Figure 3.4 shows some examples of the many possibilities. They are derived from the common amino acids and are produced by modification of the parent amino acid after the protein is synthesized by the organism in a process called posttranslational modification. Hydroxyproline and hydroxylysine differ Ifom the parent amino acids in that they have hydroxyl groups on their side chains they are found only in a few connective-tissue proteins, such as collagen. Thyroxine differs from tyrosine in that it has an extra iodine-containing aromatic group on the side... 

Hydroxyproline is formed from proline, an amino acid for which there are four codons, by posttranslational modification of the collagen precursor. 

Contains an unusual ring to the N-end amine group, which forces the CO-NH amide sequence into a fixed conformation. Can disrupt protein folding structures like a-hetix or P sheet, forcing the desired kink in the protein chain. Common in collagen, where it often undergoes a posttranslational modification to hydroxyproline. Uncommon elsewhere. 

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