Collagen proline hydroxylation

Hobza, P., j. Hurych, and R. Zahradnik Quantum Chemical Study of the Mechanism of Collagen Proline Hydroxylation. Biochim. Biophys. Acta 304, 466 (1973). 

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... 

The hydroxylation of specific Pro residues in procollagen, the precursor of collagen, requires the action of the enzyme prolyl 4-hydroxylase. This enzyme (Mt 240,000) is an a2/32 tetramer in all vertebrate sources. The proline-hydroxylating activity is found in the a subunits. (Researchers were surprised to find that the )3 subunits are identical to the enzyme protein disulfide isomerase (PDI p. 152) these subunits do not participate in the prolyl hydroxylation activity.) Each a subunit contains one atom of nonheme iron (Fe2+), and the enzyme is one of a class of hydroxylases that require a-ketoglutarate in their reactions. 

Ascorbic acid (vitamin C fig. 10.16) is the reducing agent required to maintain the activity of a number of enzymes, most notably proline hydroxylase, which forms 4-hydroxyproline residues in collagen. Hydroxyproline (see fig. 10.16c) is not synthesized biologically as a free amino acid but rather is created by modification of proline residues already incorporated into collagen. The hydroxylation reaction occurs as the protein is synthesized in the endoplasmic reticulum. At least a third of the numerous proline residues in collagen are modified in this way, substantially increasing the resistance of the protein to thermal denaturation. 

Vitagliano, L., Berisio, R., Mazzarella, L., and Zagari, A. (2001). Structural bases of collagen stabilization induced by proline hydroxylation. Biopolymers 58, 459-464. 

There are 20 different amino acids used in the synthesis of proteins these amino acids are listed in Table 3.1, which also contains the two commonly used symbols for each amino acid. The three-letter symbols are easier to remember, but the single-letter symbols are often used in writing long sequences. In many proteins some of the amino acids are modified after incorporation into proteins e.g., in collagen, a hydroxyl group is added to each of several proline residues to yield hydroxyproline residues. With the exception of proline, the a-amino acids that are incorporated into proteins can be represented by the formula shown in Fig. 3-1. 

C. Proline and lysine residues in collagen are hydroxylated in a reaction that requires vitamin C. 

No. In many proteins some of the amino acids are modified after their incorporation e.g., in collagen a hydroxyl group is added to each of several proline residues to yield hydroxyproline, and this does not constitute direct genetic encoding. 

Early in our studies it was expected that the post-translational modification of proline hydroxylation, so important to proper collagen structure and function, would raise the value of the temperature, T, for the onset of the inverse temperature transition for models of elastin. Accordingly, hydroxyproline (Hyp) was incorporated by chemical synthesis into the basic repeating sequence to give the protein-based polymers poly[fvs,i(Val-Pro-Gly-Val-Gly), fHyp( al-Hyp-Gly-Val-Gly)], where f sl -i- fnyp = 1 and values of fnyp were 0, 0.01, and 0.1. The effect of prolyl hydroxylation is shown in Figure 7.49. Replacement of proline by hydroxyproline markedly raises the temperature for hydrophobic association. Prolyl hydroxylation moves the movable cusp of... 

Chojkier, M., Spanheimer, R., and Peterkofsky, B., 1984, Specifically decreased collagen biosynthesis in scurvy dissociated from an effect on proline hydroxylation and correlated with body weight loss In vitro studies in guinea pig calvarial bones, J. Clin. Invest. 72 826-835. 

Table 9.16 shows the incorporation of [ H proline into hydroxyproline in collagen formed by granuloma tissue from control and vitamin C-defIcient (scorbutic) guinea pigs, and Table 9.17 the effects of various inhibitors of transcription and translation on the stimulation by ascorbate of proline hydroxylation in mouse fibroblasts in culture. 

A requirement for ascorbic acid in collagen formation has been well documented. This, it has been suggested, results from a utilization of this vitamin in the formation of hydroxjrproline from bound proline in a precollagen peptide (137,138) or for formation of an activated hydroxyproline (139). Mitoma and Smith (140) showed, however, that while ascorbic acid deficiency impaired collagen synthesis hydroxylation of proline was not affected. 

The proline residues on collagen are hydroxylated to increase hydrogen bonding interactions, o o... 

Since 1973, several authors have proved that there is a relationship between thermostability of collagen and the extent of hydroxylation of the proline residues31,34). Equilibrium measurements of the peptides al-CB 2 of rat tail and rat skin revealed a higher rm, for al-CB 2 (rat skin)157). The sequence of both peptides is identical except that in the peptide obtained from rat skin, the hydroxylation of the proline residues in position 3 has occurred to a higher extent than in the case of al-CB 2 (rat tail). Thus, a mere difference of 1.8 hydroxy residues per chain causes a ATm of 26 K. Obviously, there are different stabilizing interactions in the triple-helical state, that means al-CB 2 (rat skin) forms more exothermic bonds than al-CB 2 (rat tail) in the coil triple-helix transition. This leads to an additional gain of enthalpy which overcompensates the meanwhile occurring losses of entropy. 

Very few post-translational modifications have been found on tropoelastin. However, hydroxylation of 25% of the proline residues is observed [10]. The enzymatic modification of proline to hydroxyproline (Hyp) is performed by prolyl hydroxylase [11]. The purpose of this hydroxylation remains unclear and it is even proposed that Hyps in tropoelastin are a by-product of collagen hydroxylation as this occurs in the same cellular compartment [8]. 

C Ascorbic acid Coenzyme in hydroxylation of proline and lysine in collagen synthesis antioxidant enhances absorption of iron Scurvy—impaired wound healing, loss of dental cement, subcutaneous hemorrhage... 

A number of iron-containing, ascorbate-requiring hydroxylases share a common reaction mechanism in which hydroxylation of the substrate is linked to decarboxylation of a-ketoglutarate (Figure 28-11). Many of these enzymes are involved in the modification of precursor proteins. Proline and lysine hydroxylases are required for the postsynthetic modification of procollagen to collagen, and prohne hydroxylase is also required in formation of osteocalcin and the Clq component of complement. Aspartate P-hydroxylase is required for the postsynthetic modification of the precursor of protein C, the vitamin K-dependent protease which hydrolyzes activated factor V in the blood clotting cascade. TrimethyUysine and y-butyrobetaine hydroxylases are required for the synthesis of carnitine. 

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