Literature summary for 3.1.1.117 extracted from

Charavgi, M.D.; Dimarogona, M.; Topakas, E.; Christakopoulos, P.; Chrysina, E.D.
The structure of a novel glucuronoyl esterase from Myceliophthora thermophila gives new insights into its role as a potential biocatalyst (2013), Acta Crystallogr. Sect. D, 69, 63-73 .

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Cloned(Commentary)

Cloned (Comment)	Organism
gene ge2, sequence comparisons, recombinant expression of His-tagged wild-type and mutant enzymes in Pichia pastoris	Thermothelomyces thermophilus

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified unliganded wild-type and mutant S213A enzymes, and S213A mutant in complex with a substrate analogue methyl 4-O-methyl-beta-D-glucopyranuronate, sitting drop vapour diffusion method, mixing of 10 mg/ml protein in 20 mM Tris-HCl, pH 8.0, with 30% w/v PEG 3350, 0.1 M Tris-HCl, pH 8.0, for the mutant and 0.1 M sodium acetate, pH 4.6, 25% v/v PEG 550 MME for the wild-type as precipitant solutions, at 16°C, X-ray diffraction structure determination and analysis at 1.55 A, 1.9 A, and 2.35 A resolution, respectively	Thermothelomyces thermophilus
wild-type and mutant S213A, to 1.55 A and 1.9 A resolution, and mutant S213A in complex with substrate analogue, methyl 4-O-methyl-beta-D-glucopyranuronate. Serine-type hydrolase, its overall architecture follows the three-layer alphabetaalpha-sandwich hydrolase fold with a Rossmann-fold topology. Ser213, His346 and Glu236 are putative catalytic triad residues	Thermothelomyces thermophilus

Crystallization (Comment)

Organism

purified unliganded wild-type and mutant S213A enzymes, and S213A mutant in complex with a substrate analogue methyl 4-O-methyl-beta-D-glucopyranuronate, sitting drop vapour diffusion method, mixing of 10 mg/ml protein in 20 mM Tris-HCl, pH 8.0, with 30% w/v PEG 3350, 0.1 M Tris-HCl, pH 8.0, for the mutant and 0.1 M sodium acetate, pH 4.6, 25% v/v PEG 550 MME for the wild-type as precipitant solutions, at 16°C, X-ray diffraction structure determination and analysis at 1.55 A, 1.9 A, and 2.35 A resolution, respectively

Thermothelomyces thermophilus

wild-type and mutant S213A, to 1.55 A and 1.9 A resolution, and mutant S213A in complex with substrate analogue, methyl 4-O-methyl-beta-D-glucopyranuronate. Serine-type hydrolase, its overall architecture follows the three-layer alphabetaalpha-sandwich hydrolase fold with a Rossmann-fold topology. Ser213, His346 and Glu236 are putative catalytic triad residues

Thermothelomyces thermophilus

Protein Variants

Protein Variants	Comment	Organism
S213A	mutation in putative catalytic triad residue, crystallization data	Thermothelomyces thermophilus
S213A	site-directed mutagenesis, the S213A mutant exhibits a complete loss of enzyme activity towards methyl 4-O-methyl-D-glucopyranuronate, inactive mutant	Thermothelomyces thermophilus

Organism

Organism	UniProt	Comment	Textmining
Thermothelomyces thermophilus	G2QJR6	Sporotrichum thermophile	-
Thermothelomyces thermophilus	G2QJR6	member of Carbohydrate Esterase Family 15	-
Thermothelomyces thermophilus ATCC 42464	G2QJR6	Sporotrichum thermophile	-
Thermothelomyces thermophilus DSM 1799	G2QJR6	member of Carbohydrate Esterase Family 15	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His-tagged wild-type and mutant enzymes from Pichia pastoris by nickel affinity chromatography	Thermothelomyces thermophilus

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
methyl 4-O-methyl-D-glucopyranuronate + H2O	-	Thermothelomyces thermophilus	methanol + 4-O-methyl-D-glucopyranuronate	-	?
methyl 4-O-methyl-D-glucopyranuronate + H2O	-	Thermothelomyces thermophilus ATCC 42464	methanol + 4-O-methyl-D-glucopyranuronate	-	?
methyl 4-O-methyl-D-glucopyranuronate + H2O	-	Thermothelomyces thermophilus DSM 1799	methanol + 4-O-methyl-D-glucopyranuronate	-	?
methyl-4-O-methyl-D-glucuronate + H2O	-	Thermothelomyces thermophilus	methanol + 4-O-methyl-D-glucuronate	-	?
methyl-4-O-methyl-D-glucuronate + H2O	-	Thermothelomyces thermophilus ATCC 42464	methanol + 4-O-methyl-D-glucuronate	-	?
methyl-4-O-methyl-D-glucuronate + H2O	-	Thermothelomyces thermophilus DSM 1799	methanol + 4-O-methyl-D-glucuronate	-	?

Subunits

Subunits	Comment	Organism
?	x * 43000, recombinant His-tagged enzyme, SDS-PAGE	Thermothelomyces thermophilus
More	the three-dimensional protein structures of wild-type and mutant enzymes have an alpha/beta-hydrolase fold with a three-layer alphabetaalpha-sandwich architecture and a Rossmann topology and comprise one molecule per asymmetric unit	Thermothelomyces thermophilus

Synonyms

Synonyms	Comment	Organism
4-O-methyl-glucuronoyl methylesterase	-	Thermothelomyces thermophilus
Ge2	-	Thermothelomyces thermophilus
glucuronoyl esterase	-	Thermothelomyces thermophilus
StGE2	-	Thermothelomyces thermophilus

General Information

General Information	Comment	Organism
evolution	the enzyme is a member of the CE15 family of carbohydrate esterases	Thermothelomyces thermophilus
additional information	three-dimensional homology structure modelling of StGE2 by molecular replacement using the structure of Cip2_GE (PDB ID 3pic, UniProt ID G0RV93) from Hypocrea jecorina strain QM6a as a starting model. The three-dimensional protein structures of wild-type and mutant enzymes have an alpha/beta-hydrolase fold with a three-layer alphabetaalpha-sandwich architecture and a Rossmann topology and comprise one molecule per asymmetric unit. The residues lining the H12 alpha-helix and the preceding loop region are distorted compared with those in the Cip2_GE structure since the N-linked glycosylation sequence motif (Asn-X-Ser) and the N-acetylglucosamine molecule bound at Asn447 observed in the latter are missing in StGE2 (Asn-X-Ala) is the corresponding motif in StGE2. Determination of the StGE2 catalytic site structure, overview. The catalytic triad residues, namely Ser213, Glu236 and His346, participate in a concrete ready-for-nucleophilic-attack configuration	Thermothelomyces thermophilus