| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
|
Plant Physiology 134:1388-1400 (2004) © 2004 American Society of Plant Biologists The Crystal Structures of Zea mays and Arabidopsis 4-Hydroxyphenylpyruvate DioxygenaseMax-Planck-Institut für Biochemie, Abteilung für Strukturforschung, 82152 Martinsried, Germany
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 60° 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.
Dioxygenases play a key role in the degradation of aromatic compounds. 4-hydroxyphenylpyruvate dioxygenase (HPPD; EC 1.13.11.27) is an important enzyme in both Tyr and Phe catabolism of most organisms and in the biosynthesis of plastoquinones and tocopherols in plants that starts with the reaction product homogentisate. HPPD catalyzes a complex reaction involving oxidative decarboxylation of the 2-keto acid side chain of 4-hydroxyphenylpyruvate (HPP), accompanied by hydroxylation of the aromatic ring, 1,2-migration of the carboxymethyl group, and consumption of one molecule of dioxygen (Jefford and Cadby, 1981  ; Fig. 1A
). Steady state kinetic experiments indicate an ordered bi bi mechanism in which HPP is the first substrate to bind and carbon dioxide is the first product to dissociate (Rundgren, 1977). Therefore, it seems that HPP is an activating effector for the reaction of HPPD with molecular oxygen, leading to formation of the first oxygenated intermediate (Johnson-Winters et al., 2003). Similar mechanisms have been proposed for other Fe2+-dependent oxygenases (Arciero et al., 1985; Harpel and Lipscomb, 1990; Brown et al., 1995; Shu et al., 1995; Roach et al., 1997; Solomon et al., 2000). The shift of the decarboxylated side chain is still poorly understood but the migration of smaller substituents is observed in other aromatic oxygenases (Sono et al., 1996) and may involve an arene oxide intermediate (Forbes and Hamilton, 1994).
HPPD belongs to the  -keto acid-dependent group of dioxygenases; however, in contrast to other enzymes of this class, HPPD, catalyzes the incorporation of both atoms of molecular oxygen into a single substrate. Moreover, the typical -keto acid cosubstrate, -ketoglutarate, such as in prolyl hydroxylase, cephalosporin synthase, and clavaminate synthase, forms part of the substrate HPP (Que and Ho, 1996; Solomon et al., 2000). Therefore, it is not surprising that the sequences of all HPPD genes available to date reveal only low homology to other -keto acid dependent dioxygenases (Prescott, 1993). In addition, the HPPDs from prokaryotes and eukaryotes show a limited sequence identity of only about 30%, which is also reflected by their different oligomerization state. All studied mammalian HPPDs behave as homodimers of 43- to 49-kD subunits (Wada et al., 1975; Lindblad et al., 1977b; Roche et al., 1982; Endo et al., 1992; Rüetschi et al., 1993), whereas the Pseudomonas fluorescens enzyme is a homotetramer of 41-kD subunits (Lindstedt and Odelhög, 1987) as confirmed by structural analysis (Serre et al., 1999). Nevertheless, all HPPD enzymes purified and studied so far contain nonheme Fe2+ as an essential cofactor (Wada et al., 1975; Lindblad et al., 1977b; Roche et al., 1982; Endo et al., 1992; Rüetschi et al., 1993; Kim and Petersen, 2002; Johnson-Winters et al., 2003) with a mononuclear iron binding motif in the C-terminal half of the protein in which the Fe2+ ion is coordinated by two His and one carboxylic acid residues.
Due to its central role in the metabolism of aromatic amino acids in mammals and quinone synthesis in plants, the inhibition of HPPD has recently become the focus of considerable research interest. In plant tissues the HPPD reaction product homogentisate (2,5-dihydroxyphenylacetate) is the aromatic precursor of plastoquinones and tocopherols, two major classes of lipid-soluble quinone compounds in higher plant chloroplasts. The first is known for its role as an electron carrier between PSII and the cytochrome b6/f complex and as an electron carrier for NAD(P)H-plastoquinone oxidoreductase (Berger et al., 1993
Defects of Tyr catabolism in humans range from relatively mild symptoms in type II tyrosinemia (Huhn et al., 1998
Different classes of HPPD inhibitors have been developed including sulcotrione, mesotrione, and 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione (NTBC) (Fig. 1) that are based on a triketone backbone and may mimic a reaction coordinate transition state or intermediate (Kavana and Moran, 2003
To date, HPPDs of only a few plant sources have been sequenced and partially cloned, including the enzymes of Coleus blumei, Hordeum vulgare, Daucus carota, and Arabidopsis (Bartley et al., 1997 Here, we present the first crystal structures of plant HPPDs from Zea mays and Arabidopsis at a resolution of 2.0 and 3.0 Å, respectively, which reveal the dimeric architecture of the plant enzymes. As an enzyme from an anabolic pathway, HPPD is important for plant growth and therefore an interesting target for the development of new inhibitors used as herbicides.
Cloning and Expression of Plant HPPDs
Plant HPPD enzymes differ significantly at their N termini from both the bacterial and mammalian enzymes as they have extensions of at least 30 amino acids. These residues may serve as subcellular targeting signals, e.g. chloroplast import signal as HPPD could contribute to both anabolic synthesis of prenylquinones in chloroplasts and to catabolism of aromatic amino acids in the cytosol (Fiedler et al., 1982
In order to determine the shortest protein construct for crystallography we purified the native protein from Z. mays seedlings grown for 6 d. Protein sequencing yielded Ala-18 as native N terminus. In contrast, the first codon of the cloned cDNA encoded Phe-34 obtained with primer sequences based on the consensus sequence of Oryza sativa expressed sequence tags, H. vulgare, D. carota, and Arabidopsis (Linden, 2000 To clone Arabidopsis HPPD, total mRNA was isolated from seedlings. Primer sequences for cDNA synthesis were deduced from the deposited gene sequence of Bartley (1997). The amplified DNA was cloned in pET14b. Arabidopsis HPPD could be expressed as soluble protein in Escherichia coli under less stringent conditions than Z. mays HPPD. The N-terminal His-tag was cleaved off for structural studies and the recombinant protein comprised residues Gly-2 to Gly-445 preceded by Gly-Ser-His-Met from the cleavage site.
HPPDs from Z. mays and Arabidopsis have been crystallized in the orthorhombic space group P212121 and the tetragonal space group P41212, respectively. Crystals diffracted to 2.0 Å and 3.0 Å, respectively (Table I). The structure of Z. mays HPPD has been solved by single isomorphous replacement and 4-fold noncrystallographic symmetry averaging and refined at 2.0 Å resolution (Table I). The model comprises residues Phe-37 to Leu-431 for all four asymmetric monomers, one iron atom per monomer, and 847 solvent molecules. In addition, monomer A shows density for Arg-36, monomers B and C also for Phe-34 at the N terminus, whereas Glu-432 is visible in monomer B, leaving the C-terminal residues Ala-433 to Gln-435 disordered in all monomers. No electron density could be found for the N-terminal residues Ala-18 to Asn-33 and residues Glu-249 to Ser-256 in all four monomers, indicating their high flexibility. Residues Lys-397 to Glu-403 located in the insertion including
The structure of HPPD from Arabidopsis was solved by the molecular replacement method using the physiological dimer of the Z. mays structure and refined at 3.0 Å resolution. The model of Arabidopsis HPPD is composed of residues Val-33 to Phe-428 in monomer A and Val-33 to Thr-437 in monomer B. The iron ions were clearly visible in the Fo-Fc difference electron density map. There was no electron density for residues Gly-2 to Phe-32 and residues located in loop structures including Leu-107 to Thr-116, Ala-194 to Glu-201, Asp-211 to Phe-215 in both molecules, for residues Glu-252 to Glu-262 in monomer A, and Ala-255 to Glu-262 in molecule B.
The Ramachandran plot (Ramachandran and Sasisekharan, 1968
The asymmetric unit of the Z. mays HPPD crystals of space group P212121 contains two dimers of 44.8-kD subunits (Fig. 2A
). Crystals of Arabidopsis HPPD of space group P41212 contain one dimer per asymmetric unit. As both the Z. mays and Arabidopsis dimers show an identical dimerization mode we conclude that these dimers also reflect the physiological oligomeric state present in solution. The inspection of lattice contacts indicates no formation of higher oligomers in the crystals. For the Arabidopsis enzyme gel permeation chromatography and dynamic light scattering showed a Mr of 90 kD and 102 kD, respectively, in accordance with a calculated Mr of 97.6 kD (Linden, 2000
As both plant structures are quite similar to each other we will confine the description to the Z. mays HPPD. Figure 2 shows ribbon diagrams of the Z. mays monomer and dimer, and a structure based sequence alignment and assignment of secondary structure elements of the plant and bacterial enzymes is provided in Figure 3 .
The structures reveal that the monomers of both enzymes are folded into two structural domains that are arranged as an N- and C-terminal open  -barrel of eight -strands each. The N-terminal domain (residues Arg-36 to Asp-211 in Z. mays and residues Val-33 to Asp-218 in Arabidopsis) has apparently no direct catalytic function, whereas the C-terminal domain (Tyr-212 to Glu-432 in Z. mays and Tyr-219 to Thr-437 in Arabidopsis) harbors the iron binding site (His-219, His-301, and Glu-387) in the center of the -barrel, the surrounding catalytic residues, and the -helix H11 that probably functions as a gate controlling substrate access to the active site.
Both barrels pack against each other related by a pseudo-2-fold axis and form an extended
The monomer of the bacterial HPPD from P. fluorescens shares the overall fold with plant HPPDs. The superposition of the C
These deviations mainly relate to the different oligomerization modes (see below) and include two prominent insertions into the N-terminal domain from Ser-81 to Ala-86 and Ala-106 to Ser-120 of six and 15 residues, respectively. Interestingly, the Arabidopsis HPPD has a longer insertion of 21 residues in total at the latter position. However, the added residues in the Z. mays sequence that are located at an outer edge of the dimer contact are partly disordered. This large insertion induces a drastic shift of
In contrast to the dimeric plant enzymes, the bacterial P. fluorescens HPPD behaves as a tetramer of 222 point symmetry with the shape of a flat parallelepiped and tetramerization is dominated by N terminal-N terminal contacts with C-terminal domains interacting only with one neighboring domain (Serre et al., 1999
Surprisingly, the plant HPPDs dimerize in a completely different mode using an orthogonal molecular surface (Figs. 2A and 4
). Dimer formation buries an average of 1,355 Å2 in Arabidopsis HPPD and 1,544 Å2 in Z. mays HPPD in each monomer and involves both domains compared to 2,600 Å2 in P. fluorescens HPPD. However, the majority of contributions come from N-terminal-N-terminal domain interactions and no contacts are formed between C-terminal domains of neighboring subunits. These involve secondary structural elements (Ala-57 to Gly-71) including
It should be noted that the largest differences between bacterial and plant enzymes are associated with dimer formation. They affect the N-terminal half of helix H1, the region including Asp-79 to His-87 and Ala-103 to Gly-108 that are part of insertions and the region around Gly-202 to Ala-210 that has a strongly deviating backbone orientation (Fig. 2C). Differences in the C-terminal domain involve insertion from Ala-321 to Gly-324 and around Asp-380. As a consequence of the large insertion from Tyr-105 to Ser-120, the -sheet between has to shift up to 6.3 Å for Gly-169 compared to the topologically corresponding Met-109 in P. fluorescens. Intersubunit contacts are primarily hydrophilic and involve the side chains of Asp-58, Arg-65, Tyr-105, Asp-380, and Arg-381 that are strictly conserved among plant HPPDs and the nonconserved residues Ser-61, which can also be asparagines, and His-107, which can also be Asn or Pro. A prominent hydrophobic stacking interaction occurs between Phe-68 and its symmetry mate corresponding to Trp-66 in Arabidopsis HPPD (Glu-31 in P. fluorescens). An aromatic residue at that position is strictly conserved among plant HPPDs. This contact bears some resemblance to the stacking interaction between Tyr-167 in P. fluorescens HPPD and its symmetry mate. In addition, Met-322 at the edge of the contact surface packs into a small hydrophobic pocket formed from Pro-73 and Ala-210 of the neighboring molecule that are not well conserved. Taken together, the high homology in the dimer-forming regions and the strict conservation of key residues involved in dimer formation further support the view that the dimerization mode observed for the Z. mays and Arabidopsis HPPD is common to all plant HPPDs. The only major difference between both plant sequences is an insertion of six residues after Ala-112 that does not influence dimerization.
The C-terminal
This movement can be best described as swinging out of
The region preceding the hinge from Cys-409 to Phe-412 is almost identical in both plant enzymes but deviates slightly from the backbone position in the P. fluorescens structure, probably influenced by the insertion of the FG hairpin-loop in the plant enzymes. Nevertheless, this region is important as it is located in direct vicinity of the iron center including a hydrogen bond (2.9–3.1 Å) between the backbone carbonyl of Phe-412 and a water ligand of the iron atom. Although the metal ligands are not resolved at 3.0 Å in the Arabidopsis structure, the almost unaltered position of the corresponding Phe-419 suggests that a similar interaction with the coordination sphere of iron is maintained also in the open conformation. In the closed conformation -helix H11 contributes only hydrophobic side chains (Phe-412, Phe-417, Leu-420, Ile-424, and Tyr-427) to the active site compartment.
The active site of HPPD is located in the C-terminal domain formed by an eight-stranded highly twisted half-open
The structure of Z. mays HPPD, analyzed at 2.0 Å, clearly shows an octahedral coordination sphere that is comprised of three protein ligand atoms and three water molecules. The ligand sphere of the Arabidopsis HPPD was not resolved at 3.0 Å; however, there was clearly additional electron density at the iron atom that could not be satisfactorily explained by a single water molecule. The active site geometry of HPPD is very similar to that of the structurally related extradiol-cleaving catechol dioxygenase metapyrocatechase (Kita et al., 1999 ) and the biphenyl-cleaving extradiol dioxygenase DHBD (Han et al., 1995; Senda et al., 1996) although the latter enzymes have open active sites without a gating mechanism. All three enzymes share two His and a Glu residue as iron ligands. The previous structural analysis of the bacterial P. fluorescens HPPD solved at 2.4 Å demonstrated a larger ligand density that was interpreted as a monodentate acetate; metapyrocatechase showed a bound acetone molecule and DHBD a t-butanol, all of which originate from crystallization additives. Therefore, the coordination sphere of iron in these enzymes is described as a distorted tetrahedron or square pyramid as in DHBD. Interestingly, the acetone and t-butanol ligands are not close enough to the iron to serve as direct metal ion ligands. The substrate complex of P. fluorescens HPPD was modeled (Serre et al., 1999) based on the 2,3-dihydroxybiphenyl complex with oxidized inactive DHBD (Senda et al., 1996). The coordination sphere is likewise octahedral for the end-on dioxygen complex of BphC (Uragami et al., 2001). Similar to the Z. mays HPPD, other -keto acid dependent oxygenases like cephalosporin synthase and clavaminate synthase, which split their cosubstrate 2-oxoglutarate oxidatively, show a coordination sphere of the Fe2+ ion in an almost perfect octahedral geometry, with two His residues, one acidic residue, and three solvent molecules (Valegard et al., 1998; Zhang et al., 2000).
The iron center of HPPD is surrounded by an almost strictly conserved environment (Fig. 6B) that is clearly dominated by 21 hydrophobic residues in comparison to four hydrophilic and only three charged residues. The polar side chains within a radius of 5 to 8 Å around the iron are Gln-372, Asn-275, and Gln-300. With the exception of Phe-412 on the loop between
The crystal structures of Z. mays and Arabidopsis HPPD enzymes presented here provide for the first time to our knowledge structural insight into the plant representatives of these ubiquitous enzymes. As expected from the sequence homology, especially in the C-terminal half, the monomers of plant and bacterial enzymes share the overall three-dimensional fold but differ in their oligomeric state. We could demonstrate that the plant and the bacterial enzymes use nonoverlapping, orthogonal parts of the monomer surface for dimer and tetramer formation, respectively. Prominent insertions into the plant enzymes in the N-terminal domain that were not clear from previous sequence alignments are involved in the dimer contacts. In addition, the structures provide direct evidence for the substrate gating to the active site by the C-terminal -helix H11 and reveal its hinge-like rotation around Asn-416. Despite the dissimilar quaternary structures of plant and bacterial enzymes their active site architecture is remarkably well conserved, which very likely reflects steric requirements for catalysis in the multi-step reaction. Clearly, ligand binding studies will be necessary to elucidate details of the enzyme mechanism.
Cloning
Total Zea mays RNA was prepared from ethiolated seedlings grown for 6 d by using the RNeasy Mini Kit (Qiagen, Hilden, Germany). The first strand of the cDNA was obtained by reverse transcription with displayTHERMO-RT (Display Systems Biotech, Vista, CA) and an oligo(dT) primer according to the instructions of the manufacturer. Because of the high content of predicted secondary structure elements in the mRNA of Z. mays HPPD the reverse transcription reaction had to be performed with a thermostable reverse transcriptase. The HPPD gene coding for amino acids Phe-34 to Gln-435 was amplified by modified PCR introducing NdeI and BamHI restriction sites. The 5'-primer (5'-GGAATTCCTATGGCATCAGCAGCGG-3') was deduced from the N terminus of native purified Z. mays HPPD (Linden, 2000
Arabidopsis mRNA was isolated from seedlings, the cDNA synthesized by reverse transcription, and the gene amplified via PCR in a similar way. Primer sequences (5'-GGAATTCCTATGGGCCACCAAA-3' and 5'GCGGATCCTTATTGCTTGGCTTCA-AGG-3') were deduced from the known cDNA sequence (GenBank accession no. U89267; Bartley et al., 1997
Z. mays and Arabidopsis HPPDs were expressed in BL21(DE3) E. coli cells. All cultures were grown in the presence of 100 µg mL–1 carbenicillin. Isolated plasmid DNA of individual colonies was restricted and analyzed on SDS-PAGE to identify clones carrying the HPPD gene. Positive clones were used to inoculate 50 mL of Luria-Bertani (LB) broth. Successful expression of Z. mays HPPD could be monitored by a red-brownish coloration of the medium even without induction. The precultures were used to inoculate 1 L of LB broth and grown with vigorous shaking at 37°C to an OD600 of 0.6 to 0.8. Isopropylthio-
The gain of soluble Z. mays HPPD could be increased by direct inoculation of the preculture immediately after transformation without spreading on LB plates. The precultures were used to inoculate 1 L LB broth. When bacterial growth was equivalent to an OD600 of 2.0, the pH was lowered to 5.0 with 37% HCl, the medium cooled down to 20°C, and Cells were harvested by centrifugation at rcf = 5,005g* at 4°C for 20 min. The pellets were resuspended in 50 mM KPi, pH 8.0 and 500 mM NaCl (Arabidopsis HPPD) and 50 mM Tris-HCl, pH 8.0 (Z. mays HPPD), respectively, and the cells were disrupted by sonification (Branson, Heinemann, Schwäbisch Gmünd, Germany). The crude extracts were centrifuged at rcf = 75,398g* for 20 min to yield a cell-free supernatant. Arabidopsis HPPD was purified in two chromatographic steps. The supernatant was loaded onto a Co2+ Talon metal chelating column (Clontech, Palo Alto, CA), equilibrated with cell disruption buffer. Arabidopsis HPPD was eluted with 100 mM HEPES, pH 7.0, 200 mM NaCl, and 250 mM imidazole. The fractions containing HPPD were pooled, concentrated, and the buffer exchanged for 10 mM HEPES, pH 7.0 by ultrafiltration in Ultrafree filter devices (Millipore, Bedford, MA). The N-terminal His-tag was cut by incubation with thrombin (Novagen, Madison, WI) for 16 h at 4°C (0.5 unit thrombin/mg of protein). To remove thrombin and His-tag an anion exchange chromatography was carried out on Resource Q (Amersham-Pharmacia Biotech, Freiburg, Germany) in 10 mM HEPES, pH 7.0. Elution of the recombinant HPPD was done in a linear gradient from 0 to 300 mM NaCl. HPPD-containing fractions were pooled and the buffer was exchanged to 10 mM HEPES, pH 7.0 by ultrafiltration.
The soluble fraction with recombinant Z. mays HPPD was applied to a Co2+ affinity column (Clontech), equilibrated with 50 mM Tris, pH 8.0 and the protein was purified to almost homogeneity by elution using a linear imidazole gradient (50 mM Tris, pH 7.0, 500 mM imidazole). The protein was concentrated and a buffer exchange to 10 mM HEPES, pH 7.0 was carried out in an Ultrafree filter device (Millipore). The purity of the recombinant proteins was checked by SDS-PAGE stained with Coomassie Brilliant Blue (Sambrook et al., 1989 For crystallization the proteins were concentrated to 10 mg/mL (Z. mays) and 12 mg/mL (Arabidopsis), respectively. Crystals of HPPD were grown from active protein by the sitting drop vapor diffusion method. HPPD from Arabidopsis crystallized from 50% (v/v) 2-methyl-2,4-pentanediol and 6% (m/v) PEG200 in 25 mM sodium citrate, pH 5.6. Crystals grew within 4 to 6 weeks to a size of 200 x 200 x 300 µm3 at 20°C. Z. mays HPPD crystallized from 28% (v/v) 2-methyl-2,4-pentanediol, 4.9% (m/v) PEG 8000, 0.025% (v/v) dichlormethane, and 0.7 M cacodylate, pH 6.5. At 16°C crystals of Z. mays HPPD took 3 weeks to reach a size of 200 µm x 100 µm x 500 µm.
Inhouse diffraction data were collected on a MAR Research 345 imaging plate detector system mounted on a Rigaku RU-200 rotating anode operated at 50 mA and 100 kV with
The structure of Z. mays HPPD was solved by the SIRAS method. A heavy atom derivative was prepared by soaking a crystal at 16°C in 1 mM thiomersal (C9H9HgNaO2S) in mother liquor for 12 h. The data sets were integrated and scaled using the HKL suite (Otwinowski and Minor, 1997
The crystal structure of Arabidopsis HPPD was determined by molecular replacement with the program MOLREP (Collaborative Computational Project, Number 4, 1994
The stereochemistry of the refined structures was analyzed with the program PROCHECK (Laskowski et al., 1993 Received September 29, 2003; returned for revision December 22, 2003; accepted December 22, 2003.
1 Present address: m-phasys GmbH, Vor dem Kreuzberg 17, 72070 Tübingen, Germany. 
2 Bayer CropScience, BCS-R-TR, Building 6240, Alfred-Nobel-Str. 50, 40789 Monheim, Germany.
3 Present address: Antisense Pharma GmbH, Josef-Engert-Str. 9, 93053 Regensburg, Germany.
4 Present address: Division of Chemistry and Chemical Engineering, California Institute of Technology, Mail Code 114–96, Pasadena, CA 91125. Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.103.034082. * Corresponding author; fritze{at}biochem.mpg.de; fax +49–(89)–8578–3516.
Arciero DM, Orville AM, Lipscomb JD (1985) [17O] water and nitric-oxide binding by protocatechuate 4,5-dioxygenase and catechol 2,3-dioxygenase – evidence for binding of exogenous ligands to the active-site Fe2+ of extradiol dioxygenases. J Biol Chem 260: 14035–14044 Bartley GE, Maxwell CA, Wittenbach VA, Scolnik PA (1997) Cloning of an Arabidopsis thaliana cDNA for p-hydroxyphenylpyruvate dioxygenase. Plant Physiol 113: 1465–1468
Barton GJ (1993) ALSCRIPT: a tool to format multiple sequence alignments. Protein Eng 6: 37–40 Berger S, Ellersiek U, Westhoff P, Steinmüller K (1993) Studies on the expression of NDH-H, a subunit of the NAD(P)H-plastoquinone-oxidoreductase of higher plant chloroplasts. Planta 190: 25–31 Brown CA, Pavlosky MA, Westre TE, Zhang Y, Hedman B, Hodgson KO, Solomon EI (1995) Spectroscopic and theoretical description of the electronic structure of S=3/2 iron-nitrosyl complexes and their relation to O-2 activation by non-heme iron enzyme active sites. J Am Chem Soc 117: 715–732[CrossRef] Brünger AT, Adams PD, Clore GM, Delano WI, Gros P, Grosse-Kustleve RW, Jiang JS, Kuszewski J, Nilges N, Pannu NS, Read RJ, Rice LM, et al (1998) Crystallography and NMR systems (CNS): A new software system for macromolecular structure determination. Acta Crystallogr D 54: 905–921[CrossRef][Medline] Collaborative Computational Project, Number 4 (1994) The CCP4 suite: programs for protein crystallography. Acta Crystallogr D 50: 760–763[CrossRef][Medline] Cowtan K, Main P (1993) Improvement of macromolecular electron-density maps by the simultaneous application of real and reciprocal space constraints. Acta Crystallogr D 49: 148–157[CrossRef][Medline] Dähnhardt D, Falk J, Appel J, van der Kooij TAW, Schulz-Friedrich R, Krupinska K (2002) The hydroxyphenylpyruvate dioxygenase from Synechocystis sp. PCC 6803 is not required for plastoquinone synthesis. FEBS Lett 523: 177–181[CrossRef][ISI][Medline] Durand R, Zenk MH (1974) The homogentisate ring-cleavage pathway in the biosynthesis of acetate-derived naphthoquinones of Droseraceae. Phytochemistry 13: 1483–1492[CrossRef]
Endo F, Awata H, Tanoue A, Ishiguro M, Eda Y, Titani K, Matsuda I (1992) Primary structure deduced from complementary DNA sequence and expression in cultured cells of mammalian 4-hydroxyphenylpyruvic acid dioxygenase: evidence that the enzyme is a homodimer of identical subunits homologous to rat liver-specific alloantigen F. J Biol Chem 267: 24235–24240
Fernández-Cañón JM, Peñalva MA (1995) Molecular characterization of a gene encoding a homogentisate dioxygenase from Aspergillus nidulans and identification of its human and plant homologues. J Biol Chem 270: 21199–21205 Fiedler E, Soll J, Schultz G (1982) The formation of homogentisate in the biosynthesis of tocopherol and plastoquinone in spinach chloroplasts. Planta 155: 511–515[CrossRef][ISI] Forbes BJR, Hamilton GA (1994) Mechanism and mechanism-based inactivation of 4-hydroxyphenylpyruvate dioxygenase. Bioorg Chem 22: 343–361 Frishman D, Argos P (1995) Knowledge-based protein secondary structure assignment. Proteins 23: 566–579[CrossRef][ISI][Medline] Garcia I, Rodgers M, Lenne C, Rolland A, Sailland A, Matringe M (1997) Subcellular localization and purification of a p-hydroxyphenylpyruvate dioxygenase from cultured carrot cells and characterization of the corresponding cDNA. Biochem J 325: 761–769
Garcia I, Rodgers M, Pepin R, Hsieh TF, Matringe M (1999) Characterization and subcellular compartmentation of recombinant 4-hydroxyphenylpyruvate dioxygenase from arabidopsis in transgenic tobacco. Plant Physiol 119: 1507–1516
Han S, Eltis LD, Timmis KN, Muchmore SW, Bolin JT (1995) Crystal structure of the biphenyl-cleaving extradiol dioxygenase from a PCB-degrading pseudomonad. Science 270: 976–980
Harpel MR, Lipscomb JD (1990) Gentisate 1,2-dioxygenase from pseudomonas – substrate coordination to active-site Fe2+ and mechanism of turnover. J Biol Chem 265: 22187–22196 Huhn R, Stoermer H, Klingele B, Bausch E, Fois A, Farnetani M, Di Rocco M, Boue J, Kirk JM, Coleman R, Scherer G (1998) Novel and recurrent tyrosine aminotransferase gene mutations in tyrosinemia type II. Hum Genet 102: 305–313[CrossRef][ISI][Medline] Jefford CW, Cadby PA (1981) Evaluation of the models for the mechanism of action of 4-hydroxyphenylpyruvate dioxygenase. Experientia 37: 1134–1137[Medline] Johnson-Winters K, Purpero VM, Kavana M, Nelson T, Moran GR (2003) (4-hydroxyphenyl)pyruvate dioxygenase from Streptomyces avermitilis: the basis for ordered substrate addition. Biochemistry 42: 2072–2080[CrossRef][Medline] Kavana M, Moran GR (2003) Interaction of (4-hydroxyphenyl)pyruvate dioxygenase with the specific inhibitor 2-[2-nitro-4-(trifluoromethyl)benzoyl]-1,3-cyclohexanedione. Biochemistry 42: 10238–10245[CrossRef][Medline] Kim KH, Petersen M (2002) cDNA-cloning and functional expression of hydroxyphenylpyruvate dioxygenase from cell suspension cultures of Coleus blumei. Plant Sci 163: 1001–1009 Kita A, Kita S, Fujisama I, Inaka K, Ishida T, Horrike K, Nozaki M, Miki K (1999) An archetypical extradiol-cleaving catecholic dioxygenase: the crystal structure of catechol 2,3-dioxygenase (metapyrocatechase) from Pseudomonas putida mt-2. Structure 7: 25–34[Medline] Kleywegt GJ, Jones TA (1994) Halloween: masks and bones. In S Bailey, R Hubbard, D Waller, eds, From First Map to Final Model. Science and Engineering Research Council, Daresbury Laboratory, Warrington, United Kingdom, pp 59–66 Kraulis PJ (1991) MOLSCRIPT: a program to produce both detailed and schematic plots of protein structures. J Appl Crystallogr 24: 946–950[CrossRef] Laskowski RA, MacArthur MW, Moss DS, Thornton JM (1993) PROCHECK: a program to check the stereochemical quality of protein structures. J Appl Crystallogr 26: 283–291[CrossRef] Lenne C, Matringe M, Roland A, Sailland A, Pallett KE, Douce R (1995) Partial purification and localization of p-hydroxyphenylpyruvate dioxygenase activity from cultured carrot cells. In P Mathis, ed, Photosynthesis: From Light to Biosphere, Vol 5. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp 285–288
Lindblad B, Lindstedt S, Steen G (1977a) Enzymic defects in hereditary tyrosinemia. Proc Natl Acad Sci USA 74: 4641–4645
Lindblad B, Lindstedt G, Lindstedt S, Rundgren M (1977b) Purification and some properties of human 4-hydroxyphenylpyruvate dioxygenase. J Biol Chem 252: 5073–5084 Linden L (2000) Isolierung, Klonierung, Expression, Reinigung und Kristallisation eukaryotischer 4-Hydroxyphenylpyruvat Dioxygenasen – Röntgenstrukturanalyse von Inhibitorkomplexen der löslichen anorganischen Pyrophosphatase aus Saccharomyces cerevisiae. PhD Thesis. Technische Universität München, München, Germany
Lindstedt S, Rundgren M (1982) Blue color, metal content, and substrate binding in 4-hydroxyphenylpyruvate dioxygenase from Pseudomonas sp. strain P. J. 874. J Biol Chem 257: 11922–11931 Lindstedt S, Odelhög B (1987) 4-Hydroxyphenylpyruvate dioxygenase from Pseudomonas. Method Enzymol 142: 143–148[Medline] Maxwell CA, Scolnik PA, Wittenbach VA, Gutteridge S, inventors. December 31, 1997. Plant gene for p-hydroxyphenylpyruvate dioxygenase. International Application No. PCT/US97/11295 Merritt EA, Bacon DJ (1997) Raster3D photorealistic molecular graphics. Method Enzymol 277: 505–524[ISI][Medline] Norris SR, Barrette TR, DellaPenna D (1995) Genetic dissection of carotenoid synthesis in Arabidopsis defines plastoquinone as an essential component of phytoene desaturation. Plant Cell 7: 2139–2149[Abstract]
Norris SR, Shen XH, DellaPenna D (1998) Complementation of the Arabidopsis pds1 mutation with the gene encoding p-hydroxyphenylpyruvate dioxygenase. Plant Physiol 117: 1317–1323 Otwinowski Z (1991) Proceedings of the CCP4 study weekend. In W Wolf, PR Evans, AGW Leslie, eds, Isomorphous Replacement and Anomalous Scattering. Science and Engineering Research Council, Daresbury Laboratory, Warrington, United Kingdom, pp 80–86 Otwinowski Z, Minor W (1997) Processing of x-ray diffraction data collected in oscillation mode. Method Enzymol 276: 307–326[ISI] Pallett KE, Little JP, Sheekey M, Veerasekaran P (1998) The mode of action of isoxaflutole I. Physiological effects, metabolism, and selectivity. Pestic Biochem Phys 62: 113–124[CrossRef]
Phornphutkul C, Introne WJ, Perry MB, Bernardini I, Murphey MD, Fitzpatrick DL, Anderson PD, Huizing M, Anikster Y, Gerber LH, Gahl WA (2002) Natural history of alkaptonuria. N Engl J Med 347: 2111–2121
Prescott AG (1993) A dilemma of dioxygenases (or where biochemistry and molecular biology fail to meet). J Exp Bot 44: 849–861 Que LJ, Ho RYN (1996) Dioxygen activation by enzymes with mononuclear non-heme iron active sites. Chem Rev 96: 2607–2624[CrossRef][ISI][Medline] Ramachandran GN, Sasisekharan V (1968) Conformation of polypeptides and proteins. Adv Protein Chem 23: 283–437[Medline] Roach PL, Clifton IJ, Hensgens CM, Shibata N, Schofield CJ, Hajdu J, Baldwin JE (1997) Structure of isopenicillin N synthase complexed with substrate and the mechanism of penicillin formation. Nature 387: 827–830[CrossRef][Medline] Roche PA, Moorehead TJ, Hamilton GH (1982) Purification and properties of hog liver 4-hydroxyphenylpyruvate dioxygenase. Arch Biochem Biophys 216: 62–73[Medline] Rüetschi U, Dellsen A, Sahlin P, Stenman G, Rymo L, Lindstedt S (1993) Human 4-hydroxyphenylpyruvate dioxygenase, primary structure and chromosomal localization of the gene. Eur J Biochem 213: 1081–1089[Medline]
Rundgren M (1977) Steady-state kinetics of 4-hydroxyphenylpyruvate dioxygenase from human liver.3. J Biol Chem 252: 5094–5099 Russo P, O'Regan S (1990) Visceral pathology of hereditary tyrosinemia type I. Am J Hum Genet 47: 317–324[Medline] Russo PA, Mitchell GA, Tanguay RM (2001) Tyrosinemia: a review. Pediatr Dev Pathol 4: 212–221[CrossRef][ISI] |
TOP
ABSTRACT
RESULTS AND DISCUSSION
level at the active site of Z. mays HPPD. The active site iron shows an octahedral coordination sphere of three amino acid ligands and three water molecules. b, Superimposition of the active site structures of Z. mays (blue), Arabidopsis (green), and P. fluorescens (light brown) HPPD. The view includes residues approximately 14 Å around the catalytically active iron atom. Ile-220, Leu-274, Met-302, Leu-304, Ile-373, Leu-386, Ile-388, and Lys-414 in the front were omitted for clarity. The C-terminal gating helices are shown as coils.
= CuK