The enzymes isolated from the bacteria Pseudomonas cichorii , Pseudomonas sp. ST-24 , Rhodobacter sphaeroides and Mesorhizobium loti catalyse the epimerization of various ketoses at the C-3 position, interconverting D-fructose and D-psicose, D-tagatose and D-sorbose, D-ribulose and D-xylulose, and L-ribulose and L-xylulose. The specificity depends on the species. The enzymes from Pseudomonas cichorii and Rhodobacter sphaeroides require Mn2+ [2,3].
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SYSTEMATIC NAME
IUBMB Comments
D-tagatose 3-epimerase
The enzymes isolated from the bacteria Pseudomonas cichorii [2], Pseudomonas sp. ST-24 [1], Rhodobacter sphaeroides [3] and Mesorhizobium loti [4] catalyse the epimerization of various ketoses at the C-3 position, interconverting D-fructose and D-psicose, D-tagatose and D-sorbose, D-ribulose and D-xylulose, and L-ribulose and L-xylulose. The specificity depends on the species. The enzymes from Pseudomonas cichorii and Rhodobacter sphaeroides require Mn2+ [2,3].
D-tagatose 3-epimerase (PcDTE) has a broad substrate specificity, it efficiently catalyzes the epimerization of not only D-tagatose to D-sorbose but also D-fructose to D-psicose (D-allulose) and also recognizes the deoxy sugars as substrates. Substrate recognition by the enzyme at the 1-, 2-, and 3-positions is responsible for enzymatic activity and substrate-enzyme interactions at the 4-, 5-, and 6-positions are not essential for the catalytic reaction of the enzyme leading to the broad substrate specificity of PcDTE. 1-Deoxy sugars may bind to the catalytic site in the inhibitor-binding mode. Ligand-binding structure at the catalytic site, overview. Binding structures of 6-deoxy L-psicose, 1-deoxy-3-oxo-D-galactitol, 1-deoxy-D-tagatose, 1-deoxy L-tagatose, L-erythrulose, D-talitol, and glycerol
D-tagatose 3-epimerase (PcDTE) has a broad substrate specificity, it efficiently catalyzes the epimerization of not only D-tagatose to D-sorbose but also D-fructose to D-psicose (D-allulose) and also recognizes the deoxy sugars as substrates. Substrate recognition by the enzyme at the 1-, 2-, and 3-positions is responsible for enzymatic activity and substrate-enzyme interactions at the 4-, 5-, and 6-positions are not essential for the catalytic reaction of the enzyme leading to the broad substrate specificity of PcDTE. 1-Deoxy sugars may bind to the catalytic site in the inhibitor-binding mode. Ligand-binding structure at the catalytic site, overview. Binding structures of 6-deoxy L-psicose, 1-deoxy-3-oxo-D-galactitol, 1-deoxy-D-tagatose, 1-deoxy L-tagatose, L-erythrulose, D-talitol, and glycerol
in the active site Mn2+ is coordinated by Glu152, Asp185, His211, and Glu246 at the end of the beta-barrel.O2 and O3 of D-tagatose and/or D-fructose coordinate Mn2+
D-tagatose 3-epimerase (DTE) catalyzes epimerization between D-tagatose and D-sorbose. DTE from Pseudomonas cichorii (PcDTE) has a broad substrate specificity and efficiently catalyzes the epimerization of not only D-tagatose to D-sorbose but also D-fructose to D-psicose (D-allulose)
the hydrophobic groove that acts as an accessible surface for substrate binding is formed through the dimerization of PcDTE. The sugar-ring opening of a substrate may occur in the hydrophobic groove and also that the narrow channel of the passageway to the catalytic site allows a substrate in the linear form to pass through. Ligand-binding structure at the catalytic site, overview
the hydrophobic groove that acts as an accessible surface for substrate binding is formed through the dimerization of PcDTE. The sugar-ring opening of a substrate may occur in the hydrophobic groove and also that the narrow channel of the passageway to the catalytic site allows a substrate in the linear form to pass through. Ligand-binding structure at the catalytic site, overview
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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystal structures of the enzyme alone and in complexes with D-tagatose and D-fructose are determined at resolutions of 1.79, 2.28, and 2.06 A, respectively
recombinant enzyme mutant PcDTE_C66S in complexes with four deoxy rare sugars, 6-deoxy L-psicose, 1-deoxy 3-keto D-galactitol, 1-deoxy D-tagatose, and 1-deoxy L-tagatose, and with L-erythrulose (a sugar without groups at the 5- and 6-positions), hanging drop vapor diffusion method, mixing of 0.002 ml of 6-7 mg/ml protein in 5 mM Tris-HCl, pH 8.0, with 0.002 ml of reservoir solution containing 6.0-11.0 % w/v PEG 4000 and 100 mM CH3COONa, pH 4.6, and equilibration against 0.45 ml of reservoir solution, microgravity, X-ray diffraction structure determination and analysis at 1.59-2.3 A resolution, molecular replacement using crystal structure, PDB ID 1QUL, as a search model
crystals are obtained by the sitting-drop method at room temperature. The crystal belongs to the monoclinic space group P2(1), with unit-cell parameters a = 76.80, b = 94.92, c = 91.73 A , beta = 102.82°
site-directed mutagenesis, the enzyme mutant recognizes deoxy sugars as substrates. In PcDTE_C66S/deoxy sugar complex structures, bound ligand molecules in both the linear and ring forms are detected in the hydrophobic groove, while bound ligand molecules in the catalytic site are in the linear form
recombinant C-terminally His-tagged wild-type and mutant enzymes from Escherichia coli strain JM109 by nickel affinity chromatography and ultrafiltration