Molecular and Biochemical\nCharacterization of Olive\n4‑Hydroxyphenyl Pyruvate Dioxygenase Involved in the Biosynthesis\nof Tocopherols Present in Virgin Olive Oil

Olive\n(Olea europaea) fruit contains\nhigh amounts of tocopherols that are responsible, along with secoiridoid\nphenolic compounds, for most of the antioxidant and anti-inflammatory\nproperties of virgin olive oil. This study focuses on the molecular\nand biochemical characterization of olive 4-hydroxyphenyl pyruvate\ndioxygenase (OeHPPD) catalyzing the biosynthesis of homogentisic acid,\nwhich constitutes the phenolic residue in the tocopherol molecule.\nOeHPPD is a cytoplasmic enzyme with a molecular weight of 49.8 kDa\nand a predicted tertiary structure very similar to the <i>Arabidopsis</i> enzyme that suggests similar catalytic mechanisms. OeHPPD has an\nestimated <i>K</i><sub>cat</sub> of 75.26 s<sup>–1</sup> and catalytic efficiency (<i>K</i><sub>m</sub>/<i>K</i><sub>cat</sub>) of 0.145 μM<sup>–1</sup> s<sup>–1</sup> with 4-hydroxyphenyl pyruvate as the substrate. The\nexpression analysis in fruits from selected olive cultivars harvested\nat different ripening stages indicates that the <i>Oe</i>HPPD gene is temporally regulated and cultivar-dependent. Moreover,\nthe analysis of <i>OeHPPD</i> expression in fruits affected\nby drought stress suggests that HPPD is involved in olive environmental\nadaptation.