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Literature summary extracted from

  • Lin, J.F.; Sheih, Y.L.; Chang, T.C.; Chang, N.Y.; Chang, C.W.; Shen, C.P.; Lee, H.J.
    The interactions in the carboxyl terminus of human 4-hydroxyphenylpyruvate dioxygenase are critical to mediate the conformation of the final helix and the tail to shield the active site for catalysis (2013), PLoS ONE, 8, e69733.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.13.11.27 DNA and amino acid sequence determination and analysis, human enzyme expression in Escherichia coli Homo sapiens

Protein Variants

EC Number Protein Variants Comment Organism
1.13.11.27 E254D site-directed mutagenesis, the mutant shows 5% remaining activity compared to the wild-type enzyme Homo sapiens
1.13.11.27 additional information human 4-HPPD possesses an extended C-terminus compared to other 4-HPPD enzymes. Successive truncation of the disordered tail which follows the final alpha-helix results in no changes in the Km value for 4-HPP substrate but the kcat values are significantly reduced by ca. 25 to 50%. The different conformation of E254, R378 and Q375 in the final helix might be the cause of the activity loss Homo sapiens
1.13.11.27 Q375N site-directed mutagenesis, the mutant shows that a solvent accessible channel opens to the putative substrate binding site, suggesting this is responsible for the complete loss of activity, modeling, overview. Inactive mutant Homo sapiens
1.13.11.27 Q375N/R378K site-directed mutagenesis, inactive mutant Homo sapiens
1.13.11.27 R378K site-directed mutagenesis, the mutant shows 5% remaining activity compared to the wild-type enzyme Homo sapiens

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.13.11.27 0.07
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAL381 Homo sapiens
1.13.11.27 0.08
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, wild-type enzyme and mutant DELTAG388 Homo sapiens
1.13.11.27 0.09
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAL385 Homo sapiens
1.13.11.27 0.11
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant E254D Homo sapiens
1.13.11.27 0.12
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAG379 Homo sapiens
1.13.11.27 0.24
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant R378K Homo sapiens
1.13.11.27 0.3
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAN380 Homo sapiens

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.13.11.27 45000
-
 2 * 45000, SDS-PAGE Homo sapiens

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.13.11.27 4-hydroxyphenylpyruvate + O2 Homo sapiens the conversion of 4-hydroxyphenyl-pyruvate to homogentisate involves oxidative decarboxylation, side-chain migration and aromatic hydroxylation in a single catalytic cycle. The 4-HPPD reaction is unusual in that the 2-oxoacid and prime substrate moieties are contained within the same molecule homogentisate + CO2
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.13.11.27 Homo sapiens P32754
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.13.11.27 recombinant enzyme 15fold from Escherichia coli by anion-exchange and hydrophobic interaction chromatography, and gel filtration Homo sapiens

Source Tissue

EC Number Source Tissue Comment Organism Textmining
1.13.11.27 Hep-G2 cell
-
 Homo sapiens
-

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
1.13.11.27 0.04
-
 purified recombinant mutant R378K, pH 6.5, 37°C, homogentisate production Homo sapiens
1.13.11.27 0.06
-
 purified recombinant mutant E254D, pH 6.5, 37°C, O2 consumption Homo sapiens
1.13.11.27 0.07
-
 purified recombinant mutant R378K, pH 6.5, 37°C, O2 consumption Homo sapiens
1.13.11.27 0.2
-
 purified recombinant mutant E254D, pH 6.5, 37°C, homogentisate production Homo sapiens
1.13.11.27 2.6
-
 purified recombinant wild-type enzyme, pH 6.5, 37°C, O2 consumption Homo sapiens
1.13.11.27 2.8
-
 purified recombinant wild-type enzyme, pH 6.5, 37°C, homogentisate production Homo sapiens

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.13.11.27 4-hydroxyphenylpyruvate + O2
-
 Homo sapiens homogentisate + CO2
-
 ?
1.13.11.27 4-hydroxyphenylpyruvate + O2 the conversion of 4-hydroxyphenyl-pyruvate to homogentisate involves oxidative decarboxylation, side-chain migration and aromatic hydroxylation in a single catalytic cycle. The 4-HPPD reaction is unusual in that the 2-oxoacid and prime substrate moieties are contained within the same molecule Homo sapiens homogentisate + CO2
-
 ?

Subunits

EC Number Subunits Comment Organism
1.13.11.27 homodimer 2 * 45000, SDS-PAGE Homo sapiens
1.13.11.27 More five residues of the N-terminus in recombinant 4-HPPD are TTYSD Homo sapiens

Synonyms

EC Number Synonyms Comment Organism
1.13.11.27 4-HPPD
-
 Homo sapiens

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.13.11.27 37
-
 assay at Homo sapiens

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.13.11.27 0.06
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant E254D Homo sapiens
1.13.11.27 0.07
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant R378K Homo sapiens
1.13.11.27 0.1
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAG379 Homo sapiens
1.13.11.27 0.5
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAN380 Homo sapiens
1.13.11.27 1
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAL381 Homo sapiens
1.13.11.27 1.4
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAG388 and DELTAL385 Homo sapiens
1.13.11.27 2.2
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, wild-type enzyme Homo sapiens

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.13.11.27 6.5
-
 assay at Homo sapiens

General Information

EC Number General Information Comment Organism
1.13.11.27 evolution the enzyme belongs to the non-haem Fe(II)/2-oxoacid-dependent oxygenase superfamily, which couples the oxidative decarboxylation of a 2-oxoacid (most commonly a-ketoglutarate) to the oxidation of the prime substrate Homo sapiens
1.13.11.27 malfunction deficiency in active 4-HPPD in humans results in type III tyrosinemia, a rare autosomal recessive disorder Homo sapiens
1.13.11.27 metabolism enzyme 4-HPPD catalyzes the second step in the pathway of tyrosine catabolism, the conversion of 4-hydroxyphenyl-pyruvate to homogentisate Homo sapiens
1.13.11.27 additional information the disordered C-terminal tail of the enzyme, which is extended C-terminus compared to other 4-HPPD enzymes, plays an important role in catalysis with a critical role of residue Q375 in orientating the tail and ensuring the conformation of the terminal alpha-helix to maintain the integrity of the active site for catalysis. The active site of 4-HPPD is enclosed by a C-terminal alpha-helix which is assumed to function as a gate which controls access of substrate. Interactions provided by Q375 to hold the terminal helix and the tail in proper position are critical for isolating the active site from solvent during catalysis, enzyme structure modeling, overview Homo sapiens

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.13.11.27 0.27
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant R378K Homo sapiens
1.13.11.27 0.57
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant E254D Homo sapiens
1.13.11.27 0.9
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAG379 Homo sapiens
1.13.11.27 1.5
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAN380 Homo sapiens
1.13.11.27 14
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAL381 Homo sapiens
1.13.11.27 16
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAL385 Homo sapiens
1.13.11.27 17
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, mutant DELTAG388 Homo sapiens
1.13.11.27 29
-
 4-hydroxyphenylpyruvate pH 6.5, 37°C, wild-type enzyme Homo sapiens