Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary for 4.2.1.78 extracted from

  • Ilari, A.; Franceschini, S.; Bonamore, A.; Arenghi, F.; Botta, B.; Macone, A.; Pasquo, A.; Bellucci, L.; Boffi, A.
    Structural basis of enzymatic (S)-norcoclaurine biosynthesis (2009), J. Biol. Chem., 284, 897-904.
    View publication on PubMed
Search for more literature for 4.2.1.78

Cloned(Commentary)

Cloned (Comment) Organism
the NCS protein truncated at the first 19 amino acids, with a N-terminal MTGS sequence and a His tag at the C terminus is subcloned into the NdeI and XhoI restriction site of the vector pET22-b. Transformation of chemically competent Escherichia coli strain BL21DE3 Thalictrum flavum

Crystallization (Commentary)

Crystallization (Comment) Organism
SeMet derivate of NCS unligated (2VNE) and substrate-bound (2VQ5), by the hanging-drop vapour-diffusion method using ammonium sulfate and sodium chloride as precipitant agents, SeMet NCS: 2.7A resolution, 298K, ammoniumsulfate 1.4-1.8 M, NaCl 0.2 M, acetate buffer pH 4.0-4.5, trigonal space group P3121 with unit cell dimensions a = b = 86.31 A, c = 118.36 A, alpha = beta = 90°, gamma = 120° Thalictrum flavum

Protein Variants

Protein Variants Comment Organism
E110A site-directed mutagenesis, lower but significant activity Thalictrum flavum
K122A site-directed mutagenesis, lower but significant activity Thalictrum flavum
Y108A site-directed mutagenesis abolishes the stereoselective synthesis of (S)-norcoclaurine Thalictrum flavum
Y108F site-directed mutagenesis, reduced activity Thalictrum flavum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.33
-
 4-hydroxyphenylacetaldehyde wild-type, 37°C, 0.1 M phosphate buffer pH 7.5 Thalictrum flavum
0.36
-
 4-hydroxyphenylacetaldehyde mutant Y108F, 37°C, 0.1 M phosphate buffer pH 7.5 Thalictrum flavum
0.505
-
 4-hydroxyphenylacetaldehyde mutant E110A, 37°C, 0.1 M phosphate buffer pH 7.5 Thalictrum flavum

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
4-(2-Aminoethyl)benzene-1,2-diol + 4-hydroxyphenylacetaldehyde Thalictrum flavum
-
 (S)-Norcoclaurine + H2O
-
 ?

Organism

Organism UniProt Comment Textmining
Thalictrum flavum Q67A25
-
-

Purification (Commentary)

Purification (Comment) Organism
HisTrap Fast Flow column equilibrated with a 50 mM Tris HCl buffer at pH 7.5. A linear gradient of imidazole concentration from 0 to 0.5 M (buffered at pH 7.5) is applied Thalictrum flavum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
4-(2-Aminoethyl)benzene-1,2-diol + 4-hydroxyphenylacetaldehyde
-
 Thalictrum flavum (S)-Norcoclaurine + H2O
-
 ?

Subunits

Subunits Comment Organism
dimer
-
 Thalictrum flavum
tetramer composed of two asymmetric units, each containing a single dimer (A1-B1 or A2-B2) Thalictrum flavum

Synonyms

Synonyms Comment Organism
NCS
-
 Thalictrum flavum
norcoclaurine synthase
-
 Thalictrum flavum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.8
-
 4-hydroxyphenylacetaldehyde mutant E110A, 37°C, 0.1 M phosphate buffer pH 7.5 Thalictrum flavum
1.7
-
 4-hydroxyphenylacetaldehyde mutant Y108F, 37°C, 0.1 M phosphate buffer pH 7.5 Thalictrum flavum
4.5
-
 4-hydroxyphenylacetaldehyde wild-type, 37°C, 0.1 M phosphate buffer pH 7.5 Thalictrum flavum