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Literature summary for 2.4.2.6 extracted from

  • Perez, E.; Sanchez-Murcia, P.; Jordaan, J.; Blanco, M.; Mancheno, J.; Gago, F.; Fernandez-Lucas, J.
    Enzymatic synthesis of therapeutic nucleosides using a highly versatile purine nucleoside 2'-deoxyribosyltransferase from Trypanosoma brucei (2018), ChemCatChem, 10, 4406-4416 .
No PubMed abstract available
Search for more literature for 2.4.2.6

Application

Application Comment Organism
synthesis TbPDT is proficient in the biosynthesis of numerous therapeutic nucleosides, including didanosine, vidarabine, cladribine, fludarabine, and nelarabine. TbPDT has good potential as an industrial biocatalyst for the synthesis of a wide range of therapeutic nucleosides through an efficient and environmentally friendly methodology Trypanosoma brucei brucei

Cloned(Commentary)

Cloned (Comment) Organism
gene pdt, recombinant expression of N-terminally His6-tagged enzyme in Escherichia coli strain BL21(DE3) Trypanosoma brucei brucei

Protein Variants

Protein Variants Comment Organism
additional information the structure-guided replacement of Val11 with either Ala or Ser results in variants with 2.8fold greater activity. TbPDT is covalently immobilized on glutaraldehyde-activated magnetic microspheres. MTbPDT3 is selected as the best derivative (4200 IU/g, activity recovery of 22%), and can be easily recaptured and recycled for over 25 reactions with negligible loss of activity. TbPDT is immobilized onto magnetic microspheres, which are physically characterized by scanning electron microscopy and dynamic light scattering Trypanosoma brucei brucei
V11A site-directed mutagenesis, the mutant shows increased activity with 2'-deoxyinosine and adenine compared to wild-type enzyme Trypanosoma brucei brucei
V11S site-directed mutagenesis, the mutant shows increased activity with 2'-deoxyinosine and adenine compared to wild-type enzyme Trypanosoma brucei brucei

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information Michaelis-Menten kinetics Trypanosoma brucei brucei
0.2
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant wild-type enzyme, transglycosylation, with adenine Trypanosoma brucei brucei
0.7
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant wild-type enzyme, nucleobase release Trypanosoma brucei brucei
1.2
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11A, transglycosylation, with adenine Trypanosoma brucei brucei
1.8
-
 adenine pH 6.0, 50°C, recombinant wild-type enzyme, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
2
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11S, transglycosylation, with adenine Trypanosoma brucei brucei
2.9
-
 adenine pH 6.0, 50°C, recombinant mutant V11A, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
2.9
-
 adenine pH 6.0, 50°C, recombinant mutant V11S, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
5.1
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11A, nucleobase release Trypanosoma brucei brucei
6.8
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11S, nucleobase release Trypanosoma brucei brucei

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
39070
-
 recombinant homodimeric enzyme, analytical centrifugation Trypanosoma brucei brucei

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2-deoxy-D-ribosyl-base1 + base2 Trypanosoma brucei brucei
-
 2-deoxy-D-ribosyl-base2 + base1
-
 ?
2-deoxy-D-ribosyl-base1 + base2 Trypanosoma brucei brucei 927/4 GUTat10.1
-
 2-deoxy-D-ribosyl-base2 + base1
-
 ?

Organism

Organism UniProt Comment Textmining
Trypanosoma brucei brucei Q57VC7
-
-
Trypanosoma brucei brucei 927/4 GUTat10.1 Q57VC7
-
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
9.5
-
 purified recombinant wild-type enzyme, pH 6.0, 50°C, substrates 2'-deoxyadenosine and hypoxanthine Trypanosoma brucei brucei
10.5
-
 purified recombinant wild-type enzyme, pH 6.0, 50°C, substrates 2'-deoxyadenosine and guanine Trypanosoma brucei brucei
11
-
 purified recombinant wild-type enzyme, pH 6.0, 50°C, substrates 2'-deoxyinosine and guanine Trypanosoma brucei brucei
21.4
-
 purified recombinant wild-type enzyme, pH 6.0, 50°C, substrates 2'-deoxyguanosine and adenine Trypanosoma brucei brucei
38.5
-
 purified recombinant wild-type enzyme, pH 6.0, 50°C, substrates 2'-deoxyinosine and adenine Trypanosoma brucei brucei
39
-
 purified recombinant wild-type enzyme, pH 6.0, 50°C, substrates 2'-deoxyguanosine and hypoxanthine Trypanosoma brucei brucei
50.3
-
 purified recombinant mutant V11A, pH 6.0, 50°C, substrates 2'-deoxyguanosine and adenine Trypanosoma brucei brucei
59.9
-
 purified recombinant mutant V11S, pH 6.0, 50°C, substrates 2'-deoxyguanosine and adenine Trypanosoma brucei brucei
84.7
-
 purified recombinant mutant V11A, pH 6.0, 50°C, substrates 2'-deoxyinosine and adenine Trypanosoma brucei brucei
90.7
-
 purified recombinant mutant V11S, pH 6.0, 50°C, substrates 2'-deoxyinosine and adenine Trypanosoma brucei brucei

Storage Stability

Storage Stability Organism
-20°C, purified recombinant His-tagged enzyme, 180 days, about 23% activity remaining Trypanosoma brucei brucei
-80°C, purified recombinant His-tagged enzyme, 180 days, about 23% activity remaining Trypanosoma brucei brucei
4°C, purified recombinant His-tagged enzyme, 180 days, 85% activity remaining Trypanosoma brucei brucei

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2',3'-dideoxyadenosine + 2,6-diaminopurine
-
 Trypanosoma brucei brucei 2,6-diaminopurine-2',3'-dideoxyriboside + adenine
-
 ?
2',3'-dideoxyadenosine + 2,6-diaminopurine
-
 Trypanosoma brucei brucei 927/4 GUTat10.1 2,6-diaminopurine-2',3'-dideoxyriboside + adenine
-
 ?
2',3'-dideoxyadenosine + 2-fluoroadenine
-
 Trypanosoma brucei brucei 2-fluoro-2',3'-dideoxyadenosine + adenine
-
 ?
2',3'-dideoxyadenosine + 2-fluoroadenine
-
 Trypanosoma brucei brucei 927/4 GUTat10.1 2-fluoro-2',3'-dideoxyadenosine + adenine
-
 ?
2',3'-dideoxyadenosine + 6-chloropurine
-
 Trypanosoma brucei brucei 6-chloropurine-2',3'-dideoxyriboside + adenine
-
 ?
2',3'-dideoxyadenosine + 6-mercaptopurine low activity Trypanosoma brucei brucei 6-mercaptopurine-2',3'-dideoxyriboside + adenine
-
 ?
2',3'-dideoxyadenosine + 6-methoxyguanine
-
 Trypanosoma brucei brucei 6-methoxy-2',3'-dideoxyguanosine + adenine
-
 ?
2',3'-dideoxyadenosine + 6-methylpurine low activity Trypanosoma brucei brucei 6-methylpurine-2',3'-dideoxyriboside + adenine
-
 ?
2',3'-dideoxyinosine + 2,6-diaminopurine
-
 Trypanosoma brucei brucei 2,6-diaminopurine-2',3'dideoxyriboside + inosine
-
 ?
2',3'-dideoxyinosine + 6-chloropurine low activity Trypanosoma brucei brucei 6-chloropurine-2',3'-dideoxyriboside + inosine
-
 ?
2',3'-dideoxyinosine + 6-mercaptopurine low activity Trypanosoma brucei brucei 6-mercaptopurine-2',3'-dideoxyriboside + inosine
-
 ?
2',3'-dideoxyinosine + 6-methoxyguanine low activity Trypanosoma brucei brucei 6-methoxy-2',3'-dideoxyguanosine + inosine
-
 ?
2',3'-dideoxyinosine + 6-methylpurine low activity Trypanosoma brucei brucei 6-methylpurine-2',3'-dideoxyriboside + inosine
-
 ?
2'-deoxyadenosine + guanine
-
 Trypanosoma brucei brucei 2'-deoxyguanosine + adenine
-
 ?
2'-deoxyadenosine + hypoxanthine
-
 Trypanosoma brucei brucei 2'-deoxyinosine + adenine
-
 ?
2'-deoxyguanosine + 2,6-diaminopurine
-
 Trypanosoma brucei brucei 2,6-diaminopurine-2'deoxyriboside + guanine
-
 ?
2'-deoxyguanosine + 2-chloroadenine
-
 Trypanosoma brucei brucei 2-chloro-2'-deoxyadenosine + guanine
-
 ?
2'-deoxyguanosine + 2-fluoroadenine
-
 Trypanosoma brucei brucei 2-fluoro-2'-deoxyadenosine + guanine
-
 ?
2'-deoxyguanosine + 6-chloropurine
-
 Trypanosoma brucei brucei 6-chloropurine-2'-deoxyriboside + guanine
-
 ?
2'-deoxyguanosine + 6-mercaptopurine
-
 Trypanosoma brucei brucei 6-mercaptopurine-2'-deoxyriboside + guanine
-
 ?
2'-deoxyguanosine + 6-methoxyguanine
-
 Trypanosoma brucei brucei 6-methoxy-2'-deoxyguanosine + guanine
-
 ?
2'-deoxyguanosine + 6-methylpurine
-
 Trypanosoma brucei brucei 6-methylpurine-2'-deoxyriboside + guanine
-
 ?
2'-deoxyguanosine + adenine
-
 Trypanosoma brucei brucei 2'-deoxyadenosine + guanine
-
 ?
2'-deoxyguanosine + benzimidazole
-
 Trypanosoma brucei brucei benzimidazole 2'-deoxyriboside + guanine
-
 ?
2'-deoxyguanosine + hypoxanthine
-
 Trypanosoma brucei brucei 2'-deoxyinosine + guanine
-
 ?
2'-deoxyinosine + adenine
-
 Trypanosoma brucei brucei 2'-deoxyadenosine + hypoxanthine
-
 ?
2'-deoxyinosine + adenine
-
 Trypanosoma brucei brucei 927/4 GUTat10.1 2'-deoxyadenosine + hypoxanthine
-
 ?
2'-deoxyinosine + guanine
-
 Trypanosoma brucei brucei 2'-deoxyguanosine + hypoxanthine
-
 ?
2-deoxy-D-ribosyl-base1 + base2
-
 Trypanosoma brucei brucei 2-deoxy-D-ribosyl-base2 + base1
-
 ?
2-deoxy-D-ribosyl-base1 + base2
-
 Trypanosoma brucei brucei 927/4 GUTat10.1 2-deoxy-D-ribosyl-base2 + base1
-
 ?
2-fluoro-2'-deoxyinosine + 2,6-diaminopurine
-
 Trypanosoma brucei brucei 2,6-diaminopurine 2'-deoxyriboside + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + 2-chloroadenine
-
 Trypanosoma brucei brucei 2-chloro-2'-deoxyadenosine + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + 2-fluoroadenine
-
 Trypanosoma brucei brucei 2-fluoro-2'-deoxyadenosine + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + 6-chloropurine low activity Trypanosoma brucei brucei 6-chloropurine 2'-deoxyriboside + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + 6-mercaptopurine
-
 Trypanosoma brucei brucei 6-mercaptopurine 2'-deoxyriboside + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + 6-methoxyguanine
-
 Trypanosoma brucei brucei 6-methoxyguanine 2'-deoxyriboside + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + 6-methylpurine low activity Trypanosoma brucei brucei 6-methylpurine 2'-deoxyriboside + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + adenine
-
 Trypanosoma brucei brucei 2'-deoxyadenosine + 2-fluoroinosine
-
 ?
2-fluoro-2'-deoxyinosine + guanine
-
 Trypanosoma brucei brucei 2'-deoxyguanosine + 2-fluoroinosine
-
 ?
9-beta-D-arabinofuranosyl adenine + 2,6-diaminopurine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 2,6-diaminopurine + adenine
-
 ?
9-beta-D-arabinofuranosyl adenine + 2-chloroadenine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 2-chloroadenine + adenine
-
 ?
9-beta-D-arabinofuranosyl adenine + 2-fluoroadenine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 2-fluoroadenine + adenine
-
 ?
9-beta-D-arabinofuranosyl adenine + 6-chloropurine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 6-chloropurine + adenine
-
 ?
9-beta-D-arabinofuranosyl adenine + 6-methoxyguanine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 6-methoxyguanine + adenine
-
 ?
9-beta-D-arabinofuranosyl adenine + 6-methylpurine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 6-methylpurine + adenine
-
 ?
9-beta-D-arabinofuranosyl adenine + guanine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl guanine + adenine
-
 ?
9-beta-D-arabinofuranosyl adenine + hypoxanthine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl hypoxanthine + adenine
-
 ?
9-beta-D-arabinofuranosyl guanine + 2,6-diaminopurine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 2,6-diaminopurine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + 2-chloroadenine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 2-chloroadenine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + 2-fluoroadenine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 2-fluoroadenine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + 6-chloropurine low activity Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 6-chloropurine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + 6-mercaptopurine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 6-mercaptopurine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + 6-methoxyguanine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 6-methoxyguanine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + 6-methylpurine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl 6-methylpurine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + adenine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl adenine + guanine
-
 ?
9-beta-D-arabinofuranosyl guanine + hypoxanthine
-
 Trypanosoma brucei brucei 9-beta-D-arabinofuranosyl hypoxanthine + guanine
-
 ?
additional information no activity with theophylline, and with 2',3'-dideoxyadenosine/2-chloroadenine, or 2',3'-dideoxyinosine/2-fluoroadenine, or 9-beta-D-arabinofuranosyl adenine/6-mercaptopurine Trypanosoma brucei brucei ?
-
-
additional information no activity with theophylline, and with 2',3'-dideoxyadenosine/2-chloroadenine, or 2',3'-dideoxyinosine/2-fluoroadenine, or 9-beta-D-arabinofuranosyl adenine/6-mercaptopurine Trypanosoma brucei brucei 927/4 GUTat10.1 ?
-
-

Subunits

Subunits Comment Organism
homodimer 2 * 19000, recombinant enzyme, SDS-PAGE, 2 * 19090, sequence calculation Trypanosoma brucei brucei
More in general, a higher oligomerization state for NDTs is not essential for catalysis and/or stabilization of the active conformation. Binding energy analysis of the TbPDT dimer interface, overview Trypanosoma brucei brucei

Synonyms

Synonyms Comment Organism
nucleoside 2'-deoxyribosyltransferase
-
 Trypanosoma brucei brucei
PDT
-
 Trypanosoma brucei brucei
purine nucleoside 2'-deoxyribosyltransferase
-
 Trypanosoma brucei brucei
Tb927.5.1360 locus name Trypanosoma brucei brucei
TbPDT
-
 Trypanosoma brucei brucei

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
55
-
 recombinant enzyme Trypanosoma brucei brucei

Temperature Range [°C]

Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
30 80 activity range, recombinant enzyme, profile overview Trypanosoma brucei brucei

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
6.4
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant wild-type enzyme, transglycosylation, with adenine Trypanosoma brucei brucei
7.9
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant wild-type enzyme, nucleobase release Trypanosoma brucei brucei
10.2
-
 adenine pH 6.0, 50°C, recombinant wild-type enzyme, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
14.3
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11A, transglycosylation, with adenine Trypanosoma brucei brucei
17
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11S, transglycosylation, with adenine Trypanosoma brucei brucei
23.4
-
 adenine pH 6.0, 50°C, recombinant mutant V11A, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
23.8
-
 adenine pH 6.0, 50°C, recombinant mutant V11S, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
39.6
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11A, nucleobase release Trypanosoma brucei brucei
40.7
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11S, nucleobase release Trypanosoma brucei brucei

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6
-
 recombinant enzyme Trypanosoma brucei brucei

pH Range

pH Minimum pH Maximum Comment Organism
4 10 and below, activity range, recombinant enzyme, profile overview Trypanosoma brucei brucei

General Information

General Information Comment Organism
additional information molecular dynamics simulations. TbPDT is a homodimer which displays the typical 2'-deoxyribose binding site motif made up of two acidic residues from one subunit (Asp76 and Glu82) and one polar residue from the characteristic NLM motif in the neighboring monomer (Asn126#). The 5'-hydroxyl group of the incoming 2'-deoxynucleoside hydrogen bonds to the carboxylate of Asp76 and is hydrogen bonded by the carboxamide of Asn126#. The nucleophilic attack on C1' of the sugar is effected by the sidechain carboxylate of Glu82, which is held in position by hydrogen bonds donated by the phenol group of Tyr5 and the 3'-hydroxyl of the sugar. In the near-attack conformation one of the lone pairs of the M128# sulfur is poised to interact with the developing positive charge on the oxocarbenium ion of the transition state. The purine ring is sandwiched between the hydrophobic side chains of Val11 and Phe12, on one side, and those of Ile49 and Leu127# on the other side, whereas the nucleobase's N-3 atom is recognized by the carboxamide nitrogen of Asn53 Trypanosoma brucei brucei

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
5.67
-
 adenine pH 6.0, 50°C, recombinant wild-type enzyme, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
5.99
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11S, nucleobase release Trypanosoma brucei brucei
7.77
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11A, nucleobase release Trypanosoma brucei brucei
8.07
-
 adenine pH 6.0, 50°C, recombinant mutant V11A, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
8.21
-
 adenine pH 6.0, 50°C, recombinant mutant V11S, transglycosylation, with 2'-deoxyinosine Trypanosoma brucei brucei
8.5
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11S, transglycosylation, with adenine Trypanosoma brucei brucei
11.29
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant wild-type enzyme, nucleobase release Trypanosoma brucei brucei
11.92
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant mutant V11A, transglycosylation, with adenine Trypanosoma brucei brucei
32
-
 2'-deoxyinosine pH 6.0, 50°C, recombinant wild-type enzyme, transglycosylation, with adenine Trypanosoma brucei brucei