Literature summary for 3.8.1.5 extracted from

Fung, H.; Gadd, M.; Drury, T.; Cheung, S.; Guss, J.; Coleman, N.; Matthews, J.
Biochemical and biophysical characterisation of haloalkane dehalogenases DmrA and DmrB in Mycobacterium strain JS60 and their role in growth on haloalkanes (2015), Mol. Microbiol., 97, 439-453.

Application

Application	Comment	Organism
environmental protection	HLD-containing bacteria are interesting as a cleanup technology for toxic haloalkane wastes produced from industries such as plastics and pesticides manufacture	Mycolicibacterium rhodesiae
synthesis	the enzymes are of interest for biocatalysis, due to their ability to create enantiomerically pure alcohols	Mycolicibacterium rhodesiae

Cloned(Commentary)

Cloned (Comment)	Organism
gene dmrA, sequence comparisons and phylogenetic analysis, overexpression of C-terminally His6-tagged enzyme in Escherichia coli strain BL21 (DE3)	Mycolicibacterium rhodesiae
gene dmrB, sequence comparisons and phylogenetic analysis, overexpression of C-terminally His6-tagged enzyme in Escherichia coli strain BL21 (DE3)	Mycolicibacterium rhodesiae

Crystallization (Commentary)

Crystallization (Comment)	Organism
X-ray diffraction structure determination and analysis, hanging drop vapour diffusion method, mixing of equal volumes of 18 mg/ml protein solution and well solution containing 0.1 M Bis-Tris propane, pH 6.5, 0.2 M KSCN, 16% w/v PEG 3350, to 0.004 ml drops, equilibration against 1 ml reservori solution, microseeding, crystals of selenomethionyl-labeled DmrA diffract to 1.7 A resolution, PDB ID 4MJE	Mycolicibacterium rhodesiae
X-ray diffraction structure determination, hanging drop vapour diffusion method, mixing of equal volumes of protein solution and well solution, best diffracting crystals for DmrB are generated using a stock protein concentration of 5 mg/ml against 0.1 M MES, pH 6.5, 30% PEG 1500, crystals of DmrB diffract only to a resolution of 8 A	Mycolicibacterium rhodesiae

Crystallization (Comment)

Organism

X-ray diffraction structure determination and analysis, hanging drop vapour diffusion method, mixing of equal volumes of 18 mg/ml protein solution and well solution containing 0.1 M Bis-Tris propane, pH 6.5, 0.2 M KSCN, 16% w/v PEG 3350, to 0.004 ml drops, equilibration against 1 ml reservori solution, microseeding, crystals of selenomethionyl-labeled DmrA diffract to 1.7 A resolution, PDB ID 4MJE

Mycolicibacterium rhodesiae

X-ray diffraction structure determination, hanging drop vapour diffusion method, mixing of equal volumes of protein solution and well solution, best diffracting crystals for DmrB are generated using a stock protein concentration of 5 mg/ml against 0.1 M MES, pH 6.5, 30% PEG 1500, crystals of DmrB diffract only to a resolution of 8 A

Mycolicibacterium rhodesiae

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
additional information	-	additional information	Michaelis-Menten kinetics	Mycolicibacterium rhodesiae
1.9	-	4-bromobutyronitrile	pH 8.0, 25°C, recombinant enzyme	Mycolicibacterium rhodesiae

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
35000	-	NMR spectroscopy and gel filtration	Mycolicibacterium rhodesiae
35000	-	4 * 35000, multi-angle laser-light scattering analysis, NMR spectroscopy	Mycolicibacterium rhodesiae
150000	-	NMR spectroscopy and gel filtration	Mycolicibacterium rhodesiae

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
1-haloalkane + H2O	Mycolicibacterium rhodesiae	-	a primary alcohol + halide	-	?
1-haloalkane + H2O	Mycolicibacterium rhodesiae JS60	-	a primary alcohol + halide	-	?

Organism

Organism	UniProt	Comment	Textmining
Mycolicibacterium rhodesiae	G4I2J6	derived from a vinylchloride contaminated site, using vinylchloride as the sole carbon and energy source for enrichment and isolation, gene dmrA	-
Mycolicibacterium rhodesiae	G4I5P8	derived from a vinylchloride contaminated site, using vinylchloride as the sole carbon and energy source for enrichment and isolation, gene dmrB	-
Mycolicibacterium rhodesiae JS60	G4I2J6	derived from a vinylchloride contaminated site, using vinylchloride as the sole carbon and energy source for enrichment and isolation, gene dmrA	-
Mycolicibacterium rhodesiae JS60	G4I5P8	derived from a vinylchloride contaminated site, using vinylchloride as the sole carbon and energy source for enrichment and isolation, gene dmrB	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant C-terminally His6-tagged enzyme from Escherichia coli by nickel affinity chromatography and gel filtration	Mycolicibacterium rhodesiae
recombinant C-terminally His6-tagged enzyme from Escherichia coli by nickel affinity chromatography, anion exchange chromatography and gel filtration	Mycolicibacterium rhodesiae

Source Tissue

Source Tissue	Comment	Organism	Textmining
additional information	proteomic analysis of strain JS60	Mycolicibacterium rhodesiae	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
1,2-dibromoethane + H2O	-	Mycolicibacterium rhodesiae	2-bromoethanol + bromide	-	?
1,2-dibromoethane + H2O	high activity	Mycolicibacterium rhodesiae	2-bromoethanol + bromide	-	?
1,2-dibromoethane + H2O	high activity	Mycolicibacterium rhodesiae JS60	2-bromoethanol + bromide	-	?
1,2-dibromoethane + H2O	-	Mycolicibacterium rhodesiae JS60	2-bromoethanol + bromide	-	?
1,2-dichloroethane + H2O	low activity	Mycolicibacterium rhodesiae	2-chloroethanol + chloride	-	?
1,2-dichloroethane + H2O	low activity	Mycolicibacterium rhodesiae JS60	2-chloroethanol + chloride	-	?
1,6-dibromohexane + H2O	-	Mycolicibacterium rhodesiae	6-bromohexanol + bromide	-	?
1,6-dibromohexane + H2O	-	Mycolicibacterium rhodesiae JS60	6-bromohexanol + bromide	-	?
1-bromobutane + H2O	best substrate	Mycolicibacterium rhodesiae	1-butanol + bromide	-	?
1-bromobutane + H2O	best substrate	Mycolicibacterium rhodesiae JS60	1-butanol + bromide	-	?
1-bromohexane + H2O	-	Mycolicibacterium rhodesiae	1-hexanol + bromide	-	?
1-bromohexane + H2O	best substrate	Mycolicibacterium rhodesiae	1-hexanol + bromide	-	?
1-bromohexane + H2O	best substrate	Mycolicibacterium rhodesiae JS60	1-hexanol + bromide	-	?
1-bromopentane + H2O	-	Mycolicibacterium rhodesiae	1-pentanol + bromide	-	?
1-bromopentane + H2O	low activity	Mycolicibacterium rhodesiae	1-pentanol + bromide	-	?
1-bromopropane + H2O	-	Mycolicibacterium rhodesiae	1-propanol + bromide	-	?
1-haloalkane + H2O	-	Mycolicibacterium rhodesiae	a primary alcohol + halide	-	?
1-haloalkane + H2O	-	Mycolicibacterium rhodesiae JS60	a primary alcohol + halide	-	?
1-iodobutane + H2O	-	Mycolicibacterium rhodesiae	1-butanol + iodide	-	?
1-iodopropane + H2O	-	Mycolicibacterium rhodesiae	1-propanol + iodide	-	?
4-bromobutyronitrile + H2O	low activity	Mycolicibacterium rhodesiae	4-hydroxybutyronitrile + bromide	-	?
4-bromobutyronitrile + H2O	high activity	Mycolicibacterium rhodesiae	4-hydroxybutyronitrile + bromide	-	?
bromocyclohexane + H2O	low activity	Mycolicibacterium rhodesiae	cyclohexanol + bromide	-	?
bromocyclohexane + H2O	low activity	Mycolicibacterium rhodesiae JS60	cyclohexanol + bromide	-	?
additional information	no or poor activity with 1,1,2-trichloroethane and chloroform	Mycolicibacterium rhodesiae	?	-	?
additional information	no or poor activity with chloroform	Mycolicibacterium rhodesiae	?	-	?
additional information	no or poor activity with 1,1,2-trichloroethane and chloroform	Mycolicibacterium rhodesiae JS60	?	-	?
additional information	no or poor activity with chloroform	Mycolicibacterium rhodesiae JS60	?	-	?

Subunits

Subunits	Comment	Organism
monomer	1 * 35000, multi-angle laser-light scattering analysis, NMR spectroscopy	Mycolicibacterium rhodesiae
More	the protein contains 28% helical and 21% beta-strand content	Mycolicibacterium rhodesiae
More	the protein contains 31% helical and 19% beta-strand content	Mycolicibacterium rhodesiae
tetramer	4 * 35000, multi-angle laser-light scattering analysis, NMR spectroscopy	Mycolicibacterium rhodesiae

Synonyms

Synonyms	Comment	Organism
DmrA	-	Mycolicibacterium rhodesiae
DmrB	-	Mycolicibacterium rhodesiae
HLD	-	Mycolicibacterium rhodesiae

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Mycolicibacterium rhodesiae
38	-	with 1-bromohexane	Mycolicibacterium rhodesiae

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
3.1	-	4-bromobutyronitrile	pH 8.0, 25°C, recombinant enzyme	Mycolicibacterium rhodesiae

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8	-	assay at	Mycolicibacterium rhodesiae
8	-	with 1-bromohexane	Mycolicibacterium rhodesiae

Expression

Organism	Comment	Expression
Mycolicibacterium rhodesiae	haloalkane-mediated upregulation of dmrA is high by 23 to 33fold, the enzyme is also upregulated but to a lesser extent in response to starvation	up
Mycolicibacterium rhodesiae	haloalkane-mediated upregulation of dmrB is modest by 6 to 13fold, the enzyme is also upregulated but to a lesser extent in response to starvation	up

General Information

General Information	Comment	Organism
evolution	the enzyme is a member of the alpha/beta hydrolase superfamily, which also includes epoxide hydrolases and carboxylesterases. Comparison of active site cavities and access tunnels of HLDs of type I and type II HLDs	Mycolicibacterium rhodesiae
evolution	the enzyme is a member of the alpha/beta hydrolase superfamily, which also includes epoxide hydrolases and carboxylesterases. Comparison of active site cavities and access tunnels of HLDs of type I and type II HLDs. The major structural difference between DmrA (HLD subfamily I) and the well-studied enzymes of HLD subfamily II is the different arrangement of helices in the cap domain	Mycolicibacterium rhodesiae
physiological function	haloalkane dehalogenases enable the first step in bacterial growth on haloalkanes as carbon and energy sources	Mycolicibacterium rhodesiae