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Literature summary extracted from

  • Okai, M.; Ohtsuka, J.; Imai, L.F.; Mase, T.; Moriuchi, R.; Tsuda, M.; Nagata, K.; Nagata, Y.; Tanokura, M.
    Crystal structure and site-directed mutagenesis analyses of haloalkane dehalogenase LinB from Sphingobium sp. strain MI1205 (2013), J. Bacteriol., 195, 2642-2651.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.8.1.5 gene linB, DNA and amino acid sequence determination and analysis, genotyping and sequence comparisons Sphingobium indicum
3.8.1.5 gene linB, DNA and amino acid sequence determination and analysis, genotyping and sequence comparisons, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Sphingobium sp.

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
3.8.1.5 purified recombinant wild-type LinBMI, sitting drop vapor diffusion method, mixing of equal volumes of 0.001 ml of the protein solution and reservoir solution, containing 100mM Tris-HCl, pH 8.8-9.0, 200 mM CaCl2, and 17-19% w/v PEG 6000 for untagged enzyme and 100 mM Tris-HCl, pH 8.5, 200 mM MgCl2. and 20% w/v PEG 4000 for His-tagged enzyme, and equilibration against 0.3 ml of reservoir solution at 4°C or 20°C, X-ray diffraction structure determination and analysis, structure modeling Sphingobium sp.

Protein Variants

EC Number Protein Variants Comment Organism
3.8.1.5 H247A site-directed mutagenesis Sphingobium sp.
3.8.1.5 I253M site-directed mutagenesis Sphingobium sp.
3.8.1.5 L138I site-directed mutagenesis Sphingobium sp.
3.8.1.5 additional information mutations are constructed corresponding to the residues in LinBUT from Sphingobium japonicum strain UT26 Sphingobium sp.
3.8.1.5 T135A site-directed mutagenesis Sphingobium sp.
3.8.1.5 T81A site-directed mutagenesis Sphingobium sp.
3.8.1.5 V112A site-directed mutagenesis Sphingobium sp.
3.8.1.5 V134I site-directed mutagenesis Sphingobium sp.

Organism

EC Number Organism UniProt Comment Textmining
3.8.1.5 Sphingobium indicum
-
-
-
3.8.1.5 Sphingobium indicum UT26
-
-
-
3.8.1.5 Sphingobium sp. A4PEU6
-
-
3.8.1.5 Sphingobium sp. MI1205 A4PEU6
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.8.1.5 recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by metal affinity chromatography Sphingobium sp.

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.8.1.5 2,3,4,5,6-pentachlorocyclohexanol + H2O
-
 Sphingobium indicum 2,3,5,6-tetrachlorocyclohexane-1,4-diol + chloride
-
 ?
3.8.1.5 2,3,4,5,6-pentachlorocyclohexanol + H2O
-
 Sphingobium sp. 2,3,5,6-tetrachlorocyclohexane-1,4-diol + chloride
-
 ?
3.8.1.5 2,3,4,5,6-pentachlorocyclohexanol + H2O
-
 Sphingobium sp. MI1205 2,3,5,6-tetrachlorocyclohexane-1,4-diol + chloride
-
 ?
3.8.1.5 2,3,4,5,6-pentachlorocyclohexanol + H2O
-
 Sphingobium indicum UT26 2,3,5,6-tetrachlorocyclohexane-1,4-diol + chloride
-
 ?
3.8.1.5 beta-hexachlorocyclohexane + H2O
-
 Sphingobium indicum 2,3,4,5,6-pentachlorocyclohexanol + chloride
-
 ?
3.8.1.5 beta-hexachlorocyclohexane + H2O
-
 Sphingobium sp. 2,3,4,5,6-pentachlorocyclohexanol + chloride
-
 ?
3.8.1.5 beta-hexachlorocyclohexane + H2O
-
 Sphingobium sp. MI1205 2,3,4,5,6-pentachlorocyclohexanol + chloride
-
 ?
3.8.1.5 beta-hexachlorocyclohexane + H2O
-
 Sphingobium indicum UT26 2,3,4,5,6-pentachlorocyclohexanol + chloride
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.8.1.5 LinB
-
 Sphingobium indicum
3.8.1.5 LinB
-
 Sphingobium sp.
3.8.1.5 LinBMI
-
 Sphingobium sp.
3.8.1.5 LinBUT
-
 Sphingobium indicum

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.8.1.5 30
-
 assay at Sphingobium sp.

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.8.1.5 7.5
-
 assay at Sphingobium sp.

General Information

EC Number General Information Comment Organism
3.8.1.5 evolution the enzymes LinBUT and LinBMI, i.e. LinB from Sphingobium japonicum UT26 and Sphingobium sp. MI1205, respectively, catalyze the hydrolytic dechlorination of beta-hexachlorocyclohexane, mutational analysis and sequence comparisons, overview Sphingobium indicum
3.8.1.5 evolution the enzymes LinBUT and LinBMI, i.e. LinB from Sphingobium japonicum UT26 and Sphingobium sp. MI1205, respectively, catalyze the hydrolytic dechlorination of beta-hexachlorocyclohexane, mutational analysis and sequence comparisons, overview Sphingobium sp.
3.8.1.5 additional information enzyme ligand docking and molecular dynamics simulations, structure comparisons of wild-type and mutant enzymes, effects of different residues located near the active site on the specificity constants, overview Sphingobium sp.

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.8.1.5 0.0036
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant wild-type enzyme Sphingobium indicum
3.8.1.5 0.01
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant V112A Sphingobium sp.
3.8.1.5 0.027
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant V134I/H247A Sphingobium sp.
3.8.1.5 0.0271
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant wild-type enzyme Sphingobium indicum
3.8.1.5 0.07
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant T81A Sphingobium sp.
3.8.1.5 0.08
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant T135A Sphingobium sp.
3.8.1.5 0.08
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant V134L Sphingobium sp.
3.8.1.5 0.104
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant V134I/H247A Sphingobium sp.
3.8.1.5 0.124
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant V134L Sphingobium sp.
3.8.1.5 0.13
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant L138I Sphingobium sp.
3.8.1.5 0.14
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant I253M Sphingobium sp.
3.8.1.5 0.19 0.205 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant wild-type enzyme Sphingobium sp.
3.8.1.5 0.21
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant H247A Sphingobium sp.
3.8.1.5 0.21
-
 beta-hexachlorocyclohexane pH 7.5, 30°C, recombinant mutant I253M Sphingobium sp.
3.8.1.5 0.22
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant L138I Sphingobium sp.
3.8.1.5 0.23
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant V112A Sphingobium sp.
3.8.1.5 0.24
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant H247A Sphingobium sp.
3.8.1.5 0.42
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant T135A Sphingobium sp.
3.8.1.5 0.72 1 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant wild-type enzyme Sphingobium sp.
3.8.1.5 0.95
-
 2,3,4,5,6-pentachlorocyclohexanol pH 7.5, 30°C, recombinant mutant T81A Sphingobium sp.