Unusual α‑Carbon Hydroxylation of Proline Promotes Active-Site Maturation

The full extent of proline (Pro)
hydroxylation has yet to be established,
as it is largely unexplored in bacteria. We describe here a so far
unknown Pro hydroxylation activity which occurs in active sites of
polysaccharide deacetylases (PDAs) from bacterial pathogens, modifying
the protein backbone at the Cα atom of a Pro residue
to produce 2-hydroxyproline (2-Hyp). This process modifies with high
specificity a conserved Pro, shares with the deacetylation reaction
the same active site and one catalytic residue, and utilizes molecular
oxygen as source for the hydroxyl group oxygen of 2-Hyp. By providing
additional hydrogen-bonding capacity, the Pro→2-Hyp conversion
alters the active site and enhances significantly deacetylase activity,
probably by creating a more favorable environment for transition-state
stabilization. Our results classify this process as an active-site
“maturation”, which is highly atypical in being a protein
backbone-modifying activity, rather than a side-chain-modifying one.