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Literature summary extracted from

  • Li, Z.; Nair, S.K.
    Structural basis for specificity and flexibility in a plant 4-coumarate CoA ligase (2015), Structure, 23, 2032-2042 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
6.2.1.12 expressed in Escherichia coli Rosetta (DE3) cells Nicotiana tabacum

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
6.2.1.12 isoform 4CL2 in complex with Mg2+ and ATP or with AMP and coenzyme A or with three different hydroxycinnamate-AMP intermediates: 4-coumaroyl-AMP, caffeoyl-AMP, and feruloyl-AMP, hanging drop vapor diffusion method, using 20% (w/v) PEG 3350 and 0.2 M potassium nitrate or 4.4% (w/v) PEG 8000, 0.08 M sodium cacodylate (pH 6.5), 0.16 M calcium acetate, and 20% (v/v) glycerol or 25.5% (w/v) PEG 8000, 0.085 M sodium cacodylate (pH 6.5), 0.17 M sodium acetate, and 15% (v/v) glycerol or 25% (w/v) PEG 3350, 0.2 M ammonium acetate, and 0.1 M Tris (pH 8.5) Nicotiana tabacum

Protein Variants

EC Number Protein Variants Comment Organism
6.2.1.12 H237A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 K197A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 K441A inactive Nicotiana tabacum
6.2.1.12 K443A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 K526A inactive Nicotiana tabacum
6.2.1.12 M344A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 R435A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 T193A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 T336A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 Y239A the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum
6.2.1.12 Y239F the mutant shows reduced activity compared to the wild type enzyme Nicotiana tabacum

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
6.2.1.12 0.0009
-
 4-coumarate mutant enzyme K443A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.001
-
 caffeate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.0015
-
 4-coumarate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.0029
-
 ferulate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.0029
-
 4-coumarate mutant enzyme T193A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.0051
-
 4-coumarate mutant enzyme T336A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.0069
-
 4-coumarate mutant enzyme Y239F, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.0139
-
 4-coumarate mutant enzyme K197A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.1434
-
 4-coumarate mutant enzyme H237A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.2281
-
 4-coumarate mutant enzyme Y239A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.5207
-
 4-coumarate mutant enzyme M344A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.6149
-
 4-coumarate mutant enzyme R435A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 1.32
-
 sinapinate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
6.2.1.12 Mg2+ required Nicotiana tabacum

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
6.2.1.12 ATP + 4-coumarate + CoA Nicotiana tabacum
-
 AMP + diphosphate + 4-coumaroyl-CoA
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
6.2.1.12 Nicotiana tabacum O24146
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
6.2.1.12 metal ion column chromatography and gel filtration Nicotiana tabacum

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
6.2.1.12 ATP + 4-coumarate + CoA
-
 Nicotiana tabacum AMP + diphosphate + 4-coumaroyl-CoA
-
 ?
6.2.1.12 ATP + caffeate + CoA
-
 Nicotiana tabacum AMP + diphosphate + caffeoyl-CoA
-
 ?
6.2.1.12 ATP + ferulate + CoA
-
 Nicotiana tabacum AMP + diphosphate + feruloyl-CoA
-
 ?
6.2.1.12 ATP + sinapinate + CoA
-
 Nicotiana tabacum AMP + diphosphate + sinapoyl-CoA
-
 ?

Synonyms

EC Number Synonyms Comment Organism
6.2.1.12 4-coumarate:CoA ligase
-
 Nicotiana tabacum
6.2.1.12 4CL2 isoform Nicotiana tabacum

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
6.2.1.12 0.018
-
 sinapinate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.282
-
 4-coumarate mutant enzyme T336A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.996
-
 4-coumarate mutant enzyme T193A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 0.996
-
 4-coumarate mutant enzyme Y239A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 1.47
-
 4-coumarate mutant enzyme K443A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 1.98
-
 4-coumarate mutant enzyme H237A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 2.652
-
 caffeate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 3.036
-
 4-coumarate mutant enzyme R435A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 3.576
-
 ferulate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 3.978
-
 4-coumarate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 4.47
-
 4-coumarate mutant enzyme Y239F, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 7.11
-
 4-coumarate mutant enzyme K197A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 9.144
-
 4-coumarate mutant enzyme M344A, at pH 7.5 and 25°C Nicotiana tabacum

Cofactor

EC Number Cofactor Comment Organism Structure
6.2.1.12 ATP
-
 Nicotiana tabacum

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
6.2.1.12 0.0136
-
 sinapinate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 4.367
-
 4-coumarate mutant enzyme Y239A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 4.937
-
 4-coumarate mutant enzyme R435A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 13.81
-
 4-coumarate mutant enzyme H237A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 55.29
-
 4-coumarate mutant enzyme T336A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 175.6
-
 4-coumarate mutant enzyme M344A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 343.4
-
 4-coumarate mutant enzyme T193A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 511.5
-
 4-coumarate mutant enzyme K197A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 647.8
-
 4-coumarate mutant enzyme Y239F, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 1233
-
 ferulate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 1633
-
 4-coumarate mutant enzyme K443A, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 2652
-
 4-coumarate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum
6.2.1.12 2652
-
 caffeate wild type enzyme, at pH 7.5 and 25°C Nicotiana tabacum