The Crystal Structures of Zea mays and Arabidopsis 4-Hydroxyphenylpyruvate Dioxygenase

Abstract

The transformation of 4-hydroxyphenylpyruvate to homogentisate, catalyzed by 4-hydroxyphenylpyruvate dioxygenase (HPPD), plays an important role in degrading aromatic amino acids. As the reaction product homogentisate serves as aromatic precursor for prenylquinone synthesis in plants, the enzyme is an interesting target for herbicides. In this study we report the first x-ray structures of the plant HPPDs of Zea mays and Arabidopsis in their substrate-free form at 2.0 Å and 3.0 Å resolution, respectively. Previous biochemical characterizations have demonstrated that eukaryotic enzymes behave as homodimers in contrast to prokaryotic HPPDs, which are homotetramers. Plant and bacterial enzymes share the overall fold but use orthogonal surfaces for oligomerization. In addition, comparison of both structures provides direct evidence that the C-terminal helix gates substrate access to the active site around a nonheme ferrous iron center. In the Z. mays HPPD structure this helix packs into the active site, sequestering it completely from the solvent. In contrast, in the Arabidopsis structure this helix tilted by about 608 into the solvent and leaves the active site fully accessible. By elucidating the structure of plant HPPD enzymes we aim to provide a structural basis for the development of new herbicides.

Role of Substrate Positioning in the Catalytic Reaction of 4-Hydroxyphenylpyruvate Dioxygenase—A QM/MM Study

“…15 In the resting form of HPPD the active site iron is six-fold coordinated (2-His-1Glu facial triad and three water molecules) as revealed by X-ray crystal structures (PDB:1SP8,1SP9) and spectroscopic studies. 14,18,19 Results of steady-state kinetics measurements indicated that HPP binds to Fe(II) prior to dioxygen, while CO 2 is the first product released from the active site, similarly to other α-keto acid dependent enzymes. [20][21][22] Respail and co-workers performed QM/MM calculations to test the possible binding modes of HPP in the HPPD active site.…”

Broad 4-Hydroxyphenylpyruvate Dioxygenase Inhibitor Herbicide Tolerance in Soybean with an Optimized Enzyme and Expression Cassette

“…Early work with organelle fractions attributed most HPPD activity to the chloroplast in spinach (Spinacia oleracea; Fiedler et al, 1982) or Lemna gibba (Löffelhardt and Kindl, 1979). Organelle targeting can be conjectured from the observation that the N-terminal sequence of mature HPPD isolated from maize leaf begins at either Ala-17 (Fritze et al, 2004) or Ala-23 (Yang et al, 2004) with respect to the translated full-length gene. Two HPPD genes identified from EST libraries prepared from cotton tissue were 98.6% identical, each with a 23-amino acid sequence deemed likely to function in chloroplast targeting by analysis with the ChloroP prediction program (Moshiri et al, 2007).…”

Burkholderia cenocepacia C5424 Produces a Pigment with Antioxidant Properties Using a Homogentisate Intermediate

“…HppD has been described in humans (4,44), mouse (9), and rat (10), as well as plants (14,16), fungi (63), and prokaryotes (8,45). There is considerable interest in the HGA catabolic pathway, because HppD in plants is an important herbicide target (5,15,23) and many severe human diseases, like phenylketonuria (18); alkaptonuria (52); tyrosinemias I, II, and III; and hawkinsinuria (48), are associated with enzyme deficiencies in the catabolism of tyrosine.…”