Proline-trans-4-hydroxylase

Hydroxylation of Pipecolic Acid Using Recombinant Cells of E. coli Expressing Proline-trans-4-hydroxylase from Dactylsporangium sp. 

Figure 7. Hydroxylation of L-proline to trans-4-hydroxy-L-pro-line by proline 4-hydroxylase. The cosubstrate a-ketoglutarate is continuously supplied by the metabolism of growing E. coli W1485 (pWFHl), which served as biocatalyst.

Proline derivatives trans-4-hydroxy-L-proline (Hyp) is an important component of animal supportive and coimective tissues Free all-cis-4-hyAoxy-L-proline occurs m Santalum album and other pUmts Small quantities of 3-hydroxy-L-proline are also present m collagen. 4-Hydroxy-L-proline is formed mainly by hydroxylation of ribosome-bound peptidylpro-lyl-RNA, a reaction requiring ascorbic add and catalysed by proline hydroxylase (see Oxygenases). Flee 4-hydroxy-L-proline is formed by cyclization of y-hy-... 

Recently George Gardinale has been able to show that prolyl hydroxylase can catalyze the oxidative decarboxylation of a-ketoglutarate in the absence of a prolyl substrate,>at a lower but measurable rate. The same cofactors, Fe and ascorbate, are required and optimal conditions are the same for both. This suggests that a-ketoglutarate is the first acceptor of the oxygen and that a peroxy compound, perhaps enzyme bound persuccinate, then attacks the 4-trans position of proline to yield the intermediate shown above. Gleavage of the peroxy compound would then yield equal amounts of succinate and hydroxyproline. 

OG dependent hydroxylases have attracted the most attention for preparative biocatalysis because of their ability to catalyze the hydroxylation of amino adds. Although eukaryotic enzymes only catalyze the hydroxylation of pep-tidic substrates, homologous enzymes also display the ability to hydroxylate isolated amino acids including proline, isoleudne, asparagine, and lysine. The proline hydroxylases have attracted most attention, both because regio- and en-antiocomplementary enzymes that allow access to cis- and trans- forms of both... 

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