Characterization of two carnation petal prolyl 4 hydroxylases

Prolyl 4-hydroxylases (P4Hs) catalyze the proline hydroxylation, a major post-translational modification, of hydroxyproline-rich glycoproteins. Two carnation petal P4H cDNAs, (Dianthus caryophyllus prolyl 4-hydroxylase) DcP4H1 and DcP4H2, were identified and characterized at the gene expression and biochemical level in order to investigate their role in flower senescence. Both mRNAs showed similar patterns of expression with stable transcript abundance during senescence progression and differential tissue-specific expression with DcP4H1 and DcP4H2 strongly expressed in ovaries and stems, respectively. Recombinant DcP4H1 and DcP4H2 proteins were produced and their catalytic properties were determined. Pyridine 2,4-dicarboxylate (PDCA) was identified as a potent inhibitor of the in vitro enzyme activity of both P4Hs and used to determine whether inhibition of proline hydroxylation in petals is involved in senescence progression of cut carnation flowers. PDCA suppressed the climacteric ethylene production indicating a strong correlation between the inhibition of DcP4H1 and DcP4H2 activity in vitro by PDCA and the suppression of climacteric ethylene production in cut carnation flowers.