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

BRENDA support

Literature summary for 4.1.99.3 extracted from

  • Klar, T.; Kaiser, G.; Hennecke, U.; Carell, T.; Batschauer, A.; Essen, L.O.
    Natural and non-natural antenna chromophores in the DNA photolyase from Thermus thermophilus (2006), Chembiochem, 7, 1798-1806.
    View publication on PubMed

Application

Application Comment Organism
biotechnology it should be possible to perform wavelength tuning of the Thermus photolyase by using artificial flavin chromophores and photolyase variants whose antenna chromophore-binding sites are reengineered by molecular modeling and mutagenesis Thermus thermophilus

Cloned(Commentary)

Cloned (Comment) Organism
cloned in Escherichia coli Thermus thermophilus

Crystallization (Commentary)

Crystallization (Comment) Organism
hexagonal crystals are obtained by the hangingdrop vapor-diffusion method after four days. Coordinates and structure factors of the photolyase-chromophore complexes are deposited in the RCSB protein databank under the accession codes 2J07, 2J08, and 2J09 Thermus thermophilus

Protein Variants

Protein Variants Comment Organism
R46E R46E mutant which lacks a conserved arginine in the binding site for the antenna chromophore shows no flavin-mononucleotide and discloses an eightfold lower activity at 450 nm (blue light) wheras at 370 nm (UV-A light) its activity is only three times lower than wildtype enzyme Thermus thermophilus

Organism

Organism UniProt Comment Textmining
Thermus thermophilus P61497
-
-

Purification (Commentary)

Purification (Comment) Organism
cells are disrupted by an emulsifier, followed by heat treatment for 15 min at 65°C and centrifugation. Supernatant is purified by using a Blue Sepharose CL-6B affinity column. Recombinant photolyase elutes at a concentration of 1 M KCl in an increasing salt gradient. Fraction containing the protein are combined and concentrated by ultrafiltration. Gel filtration chromatography with Sephacryl S200-HR as a final step give an overal yield of 35 mg of purified CPD-photolyase per liter cell culture Thermus thermophilus

Source Tissue

Source Tissue Comment Organism Textmining

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
cyclobutadipyrimidine in DNA
-
Thermus thermophilus 2 pyrimidine residues in DNA
-
?

Synonyms

Synonyms Comment Organism
CPD-photolyase
-
Thermus thermophilus
DNA photolyase
-
Thermus thermophilus

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
assay at Thermus thermophilus

Cofactor

Cofactor Comment Organism Structure
7,8-didemethyl-8-hydroxy-5-deazariboflavin chromophore binding site of Thermus photolyase is reconstited also with 7,8-didemethyl-8-hydroxy-5-deazariboflavin (8-HDF). However, in the genome sequence of Thermus thermophilus it is found that the genes essential for the biosynthesis of 8-HDF are missing Thermus thermophilus
8-iodoflavin chromophore binding site of Thermus photolyase is reconstited also with a novel synthetic flavin, 8-iodoflavin Thermus thermophilus
flavin flavin-mononucleotide (FMN), crystal strucutre analysis reveals the binding of flavin mononucleotide as an antenna chromophore Thermus thermophilus