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Literature summary for 1.3.3.4 extracted from

  • Heinemann, I.U.; Diekmann, N.; Masoumi, A.; Koch, M.; Messerschmidt, A.; Jahn, M.; Jahn, D.
    Functional definition of the tobacco protoporphyrinogen IX oxidase substrate binding site (2007), Biochem. J., 402, 575-580.
    View publication on PubMedView publication on EuropePMC

Application

Application Comment Organism
medicine tobacco PPO2 represents a useful model system for the understanding of the structure–function relationship underlying detrimental human enzyme defects, Arg98, Phe392, Leu356 and Leu372 are functionally involved in substrate co-ordination within the active site Nicotiana tabacum

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli strain BL21(DE3)RIL Nicotiana tabacum
expression in Escherichia coli BL21(DE3)RIL containing the original or mutated pET32a-PPO2 plasmids Nicotiana tabacum

Crystallization (Commentary)

Crystallization (Comment) Organism
-
Nicotiana tabacum

Protein Variants

Protein Variants Comment Organism
F329E shows increased activity Nicotiana tabacum
F392E inferior binding of the substrate, but increased turnover rate than wild-type Nicotiana tabacum
F392H does not reveal detectable enzyme activity indicating an important role of Phe392 in substrate ring A stacking Nicotiana tabacum
F392H no detectable enzyme activity Nicotiana tabacum
L356N an only slightly inferior binding capacity compared with the wild-type and comparable catalytic PPO2 activities Nicotiana tabacum
L356N shows increased activity Nicotiana tabacum
L356V increased Km value, once the substrate is bound, catalysis is enhanced Nicotiana tabacum
L356V shows increased activity Nicotiana tabacum
L372N an only slightly inferior binding capacity compared with the wild-type and comparable catalytic PPO2 activities Nicotiana tabacum
L372N shows increased activity Nicotiana tabacum
L372V increased Km value, once the substrate is bound, catalysis is enhanced Nicotiana tabacum
L372V shows increased activity Nicotiana tabacum
N67R inferior substrate binding, acceptable catalytic efficiency Nicotiana tabacum
N67R shows increased activity Nicotiana tabacum
N67R/S374D almost wild-type level Km value Nicotiana tabacum
N67R/S374D shows increased activity Nicotiana tabacum
N67W does not reveal detectable enzyme activity Nicotiana tabacum
N67W/S374D significantly reduced enzyme activity Nicotiana tabacum
N67W/S374D shows increased activity, structural constellation mimicking the human disease variegate porphyria Nicotiana tabacum
N6W no detectable enzyme activity Nicotiana tabacum
R98A 8fold increased Km value, improved turnover Nicotiana tabacum
R98A shows increased activity Nicotiana tabacum
R98E inferior substrate binding, improved turnover Nicotiana tabacum
R98E shows increased activity Nicotiana tabacum
R98K improved turnover Nicotiana tabacum
R98K shows increased activity Nicotiana tabacum
S374D increased turnover rate Nicotiana tabacum
S374D shows increased activity Nicotiana tabacum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.00117
-
protoporphyrinogen-IX wild-type Nicotiana tabacum
0.00117
-
protoporphyrinogen-IX wild type enzyme PPO2, at 20°C Nicotiana tabacum
0.0014
-
protoporphyrinogen-IX N67R/S374D Nicotiana tabacum
0.0014
-
protoporphyrinogen-IX mutant enzyme N67R/S374D, at 20°C Nicotiana tabacum
0.0026
-
protoporphyrinogen-IX R98K Nicotiana tabacum
0.0026
-
protoporphyrinogen-IX mutant enzyme R98K, at 20°C Nicotiana tabacum
0.0073
-
protoporphyrinogen-IX L356V Nicotiana tabacum
0.0073
-
protoporphyrinogen-IX mutant enzyme L356V, at 20°C Nicotiana tabacum
0.0083
-
protoporphyrinogen-IX R98A Nicotiana tabacum
0.0083
-
protoporphyrinogen-IX mutant enzyme R98A, at 20°C Nicotiana tabacum
0.0109
-
protoporphyrinogen-IX S374D Nicotiana tabacum
0.0109
-
protoporphyrinogen-IX mutant enzyme S374D, at 20°C Nicotiana tabacum
0.011
-
protoporphyrinogen-IX L356N Nicotiana tabacum
0.011
-
protoporphyrinogen-IX mutant enzyme L356N, at 20°C Nicotiana tabacum
0.0112
-
protoporphyrinogen-IX F392E Nicotiana tabacum
0.0112
-
protoporphyrinogen-IX mutant enzyme F392E, at 20°C Nicotiana tabacum
0.0125
-
protoporphyrinogen-IX R98E Nicotiana tabacum
0.0125
-
protoporphyrinogen-IX mutant enzyme R98E, at 20°C Nicotiana tabacum
0.0164
-
protoporphyrinogen-IX L372N Nicotiana tabacum
0.0164
-
protoporphyrinogen-IX mutant enzyme L372N, at 20°C Nicotiana tabacum
0.021
-
protoporphyrinogen-IX N67W/S374D Nicotiana tabacum
0.021
-
protoporphyrinogen-IX mutant enzyme N67W/S374D, at 20°C Nicotiana tabacum
0.097
-
protoporphyrinogen-IX N67R Nicotiana tabacum
0.097
-
protoporphyrinogen-IX mutant enzyme N67R, at 20°C Nicotiana tabacum
0.103
-
protoporphyrinogen-IX L372V Nicotiana tabacum
0.103
-
protoporphyrinogen-IX mutant enzyme L372V, at 20°C Nicotiana tabacum

Localization

Localization Comment Organism GeneOntology No. Textmining
mitochondrion PPO2 Nicotiana tabacum 5739
-
plastid PPO1 Nicotiana tabacum 9536
-

Organism

Organism UniProt Comment Textmining
Nicotiana tabacum
-
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni-IDA resin column chromatography Nicotiana tabacum
to apparent homogeneity by Ni-IDA resin Nicotiana tabacum

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
4.27
-
-
Nicotiana tabacum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
protoporphyrinogen-IX + 3 O2
-
Nicotiana tabacum protoporphyrin-IX + 3 H2O2
-
?
protoporphyrinogen-IX + 3 O2
-
Nicotiana tabacum protoporphyrin-IX + 3 H2O2
-
ir

Synonyms

Synonyms Comment Organism
PPO1 plastidic PPO Nicotiana tabacum
PPO2
-
Nicotiana tabacum
PPO2 mitochondrial PPO Nicotiana tabacum
protoporphyrinogen IX oxidase
-
Nicotiana tabacum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
6
-
protoporphyrinogen-IX wild-type Nicotiana tabacum
6
-
protoporphyrinogen-IX wild type enzyme PPO2, at 20°C Nicotiana tabacum
7
-
protoporphyrinogen-IX L372N Nicotiana tabacum
7
-
protoporphyrinogen-IX mutant enzyme L372N, at 20°C Nicotiana tabacum
11
-
protoporphyrinogen-IX F392E Nicotiana tabacum
11
-
protoporphyrinogen-IX mutant enzyme F392E, at 20°C Nicotiana tabacum
12
-
protoporphyrinogen-IX N67W/S374D Nicotiana tabacum
12
-
protoporphyrinogen-IX mutant enzyme N67W/S374D, at 20°C Nicotiana tabacum
34
-
protoporphyrinogen-IX R98E Nicotiana tabacum
34
-
protoporphyrinogen-IX mutant enzyme R98E, at 20°C Nicotiana tabacum
37
-
protoporphyrinogen-IX R98K Nicotiana tabacum
37
-
protoporphyrinogen-IX mutant enzyme R98K, at 20°C Nicotiana tabacum
38
-
protoporphyrinogen-IX L356N Nicotiana tabacum
38
-
protoporphyrinogen-IX mutant enzyme L356N, at 20°C Nicotiana tabacum
97
-
protoporphyrinogen-IX N67R Nicotiana tabacum
97
-
protoporphyrinogen-IX mutant enzyme N67R, at 20°C Nicotiana tabacum
111
-
protoporphyrinogen-IX N67R/S374D Nicotiana tabacum
111
-
protoporphyrinogen-IX mutant enzyme N67R/S374D, at 20°C Nicotiana tabacum
208
-
protoporphyrinogen-IX mutant enzyme S374D, at 20°C Nicotiana tabacum
300
-
protoporphyrinogen-IX L356V Nicotiana tabacum
300
-
protoporphyrinogen-IX mutant enzyme L356V, at 20°C Nicotiana tabacum
365
-
protoporphyrinogen-IX R98A Nicotiana tabacum
365
-
protoporphyrinogen-IX mutant enzyme R98A, at 20°C Nicotiana tabacum
597
-
protoporphyrinogen-IX L372V Nicotiana tabacum
597
-
protoporphyrinogen-IX mutant enzyme L372V, at 20°C Nicotiana tabacum

Cofactor

Cofactor Comment Organism Structure
flavin dependent Nicotiana tabacum