Proline hydroxylation

As proposed schematically in Fig. 13.2, RNI may block PHD-activity, attenuate proline hydroxylation of HIF-la, dissociate HIF-la from pVHL with the consequence of protein stabilization based on decreased proteasomal degradation. 

Ivan, M. et al. HlFalpha targeted for VHL-mediated destruction by proline hydroxylation implications for O2 sensing. Science 2001, 292, 464-8. 

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. 

In the normal prolyl 4-hydroxylase reaction (Fig. 4a), one molecule of a-ketoglutarate and one of 02 bind to the enzyme. The a-ketoglutarate is oxidatively decarboxylated to form C02 and succinate. The remaining oxygen atom is then used to hydroxylate an appropriate Pro residue in procollagen. No ascorbate is needed in this reaction. However, prolyl 4-hydroxylase also catalyzes an oxidative decarboxylation of a-ketoglutarate that is not coupled to proline hydroxylation—and this is the reaction that requires ascorbate (Fig. 4b). During this reaction, the heme Fe2+ becomes oxidized, and the oxidized form of the enzyme is inactive—unable to hydroxylate proline. The ascorbate consumed in the reaction presumably functions to reduce the heme iron and restore enzyme activity. 

FIGURE 4 The reactions catalyzed by prolyl 4-hydroxylase, (a) The normal reaction, coupled to proline hydroxylation, which does not require ascorbate. The fate of the two oxygen atoms from 02 is shown in red. (b) The uncoupled reaction, in which a-ketoglutarate is oxidatively decarboxylated without hydroxylation of proline. Ascorbate is consumed stoichiometrically in this process as it is converted to dehydroascorbate. 

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

KaelinWG. 2005. Proline hydroxylation and gene expression. Ann Rev Biochem 74 115-128. 

The contryphan family of cyclic peptides, isolated from various species of cone snails, have the conserved sequence motif H3N" -X COD-WX PWC-NH2, where X is either Gly or absent, O is 4-trtf j--hydroxypro-line, and X is Glu, Asp, or Gin. The contryphans possess a distinctive number of PTMs that include tryptophan bromination, proline hydroxylation, glutamate carboxylation (7-carboxyglutamate), C-terminal amidation, and leucine and tryptophan, l to d, isomerization. ... 

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... 

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

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. 

Rosenbloom, J., and Cywinski, A., 1976, Inhibition of proline hydroxylation does not inhibit secretion of tropoelastin by chick aorta cells, FEBS Lett. 65 246-250. 

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. 

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...

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