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

BRENDA support

Literature summary for 2.7.7.31 extracted from

  • Chua, J.P.S.; Go, M.K.; Osothprarop, T.; Mcdonald, S.; Karabadzhak, A.G.; Yew, W.S.; Peisajovich, S.; Nirantar, S.
    Evolving a thermostable terminal deoxynucleotidyl transferase (2020), ACS Synth. Biol., 9, 1725-1735 .
    View publication on PubMed
Search for more literature for 2.7.7.31

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Bos taurus

Protein Variants

Protein Variants Comment Organism
K177N/E178G/K271E/F280L/M283K/H405P the thermostable mutant TdT3-2 shows a 10 C increased melting temperature as compared to the wild type enzyme. The mutant is capable of incorporating higher amount of ddCTP onto double stranded blunt end DNA Bos taurus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.00646
-
 2bA FAM oligo mutant enzyme K177N/E178G/K271E/F280L/M283K/H405P, at pH 7.2 and 37°C Bos taurus
0.00819
-
 2bA FAM oligo mutant enzyme K177N/E178G/K271E/F280L/M283K/H405P, at pH 7.2 and 37°C Bos taurus
0.0176
-
 2bA FAM oligo wild type enzyme, at pH 7.2 and 37°C Bos taurus

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ 10 mM used in assay conditions Bos taurus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
a 2'-deoxyribonucleoside 5'-triphosphate + DNAn Bos taurus
-
 diphosphate + DNAn+1
-
 ?

Organism

Organism UniProt Comment Textmining
Bos taurus P06526
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni-Sepharose column chromatography and Q Sepharose column chromatography Bos taurus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
a 2'-deoxyribonucleoside 5'-triphosphate + DNAn
-
 Bos taurus diphosphate + DNAn+1
-
 ?
Cy5-dCTP + 2bA FAM oligo 2bA FAM oligo is a primer with FAM at 2 bases from the 3' end (5'-CGCTTGCACAGGTGCGTT/iFluorT/CA-3') Bos taurus diphosphate + ?
-
 ?
additional information wild type enzyme is unable to incorporate bulky 3'-blocked nucleotides due to steric clashes Bos taurus ?
-
-

Synonyms

Synonyms Comment Organism
TdT
-
 Bos taurus
terminal deoxynucleotidyl transferase
-
 Bos taurus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
-
 Bos taurus

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
41 47 the melting temperature of the wild type enzyme is 41.5-45.7 C. At 47 C, the wild type enzyme is denatured within 5 min. At 58 C,the wild type enzyme is not active Bos taurus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0925
-
 2bA FAM oligo mutant enzyme K177N/E178G/K271E/F280L/M283K/H405P, at pH 7.2 and 47°C Bos taurus
0.893
-
 2bA FAM oligo wild type enzyme, at pH 7.2 and 37°C Bos taurus
1.2
-
 2bA FAM oligo mutant enzyme K177N/E178G/K271E/F280L/M283K/H405P, at pH 7.2 and 37°C Bos taurus

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.25
-
 2bA FAM oligo wild type enzyme, at pH 7.2 and 47°C Bos taurus
14.3
-
 2bA FAM oligo mutant enzyme K177N/E178G/K271E/F280L/M283K/H405P, at pH 7.2 and 47°C Bos taurus
50.7
-
 2bA FAM oligo wild type enzyme, at pH 7.2 and 37°C Bos taurus
146
-
 2bA FAM oligo mutant enzyme K177N/E178G/K271E/F280L/M283K/H405P, at pH 7.2 and 37°C Bos taurus