Literature summary for 5.2.1.1 extracted from

Chen, D.; Tang, H.; Lv, Y.; Zhang, Z.; Shen, K.; Lin, K.; Zhao, Y.L.; Wu, G.; Xu, P.
Structural and computational studies of the maleate isomerase from Pseudomonas putida S16 reveal a breathing motion wrapping the substrate inside (2013), Mol. Microbiol., 87, 1237-1244.

Cloned(Commentary)

Cloned (Comment)	Organism
overexpression of N- and C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)	Pseudomonas putida

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant wild-type alone, and mutant enzyme C200A in complex with substrate maleate, hanging-drop vapour-diffusion method, mixing of 15-20 mg/ml protein in 25 mM Tris-HCl, pH 8.0, 150 mM NaCl, and 2 mM DTT, with a reservoir solution containing 25% w/v PEG3,350 and 0.1 M Tris, pH 8.5, for the substrate-enzyme mutant complex, and 1.4 M (NH4)2SO4, 120 mM NaCl, and 0.1 M HEPES, pH 7.5, for the free wild-type enzyme, 14°C, X-ray diffraction structure determination and analysis at 2.95 A and 2.10 A resolution, respectively	Pseudomonas putida

Crystallization (Comment)

Organism

purified recombinant wild-type alone, and mutant enzyme C200A in complex with substrate maleate, hanging-drop vapour-diffusion method, mixing of 15-20 mg/ml protein in 25 mM Tris-HCl, pH 8.0, 150 mM NaCl, and 2 mM DTT, with a reservoir solution containing 25% w/v PEG3,350 and 0.1 M Tris, pH 8.5, for the substrate-enzyme mutant complex, and 1.4 M (NH4)2SO4, 120 mM NaCl, and 0.1 M HEPES, pH 7.5, for the free wild-type enzyme, 14°C, X-ray diffraction structure determination and analysis at 2.95 A and 2.10 A resolution, respectively

Pseudomonas putida

Protein Variants

Protein Variants	Comment	Organism
C200A	site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme, substrate-binding structure: maleate can be found to exist inside a cavity of the enzyme, totally surrounded by the protein	Pseudomonas putida
C84A	site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme	Pseudomonas putida

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
Maleate	Pseudomonas putida	-	Fumarate	-	r
Maleate	Pseudomonas putida S16	-	Fumarate	-	r

Organism

Organism	UniProt	Comment	Textmining
Pseudomonas putida	F8G0M3	-	-
Pseudomonas putida S16	F8G0M3	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant N- and C-terminally His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration	Pseudomonas putida

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
Maleate	-	Pseudomonas putida	Fumarate	-	r
Maleate	Asn17 and Asn169 play critical roles in recognizing maleate	Pseudomonas putida	Fumarate	-	r
Maleate	-	Pseudomonas putida S16	Fumarate	-	r
Maleate	Asn17 and Asn169 play critical roles in recognizing maleate	Pseudomonas putida S16	Fumarate	-	r

Synonyms

Synonyms	Comment	Organism
Pp-Iso	-	Pseudomonas putida

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
30	-	assay at	Pseudomonas putida

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8.4	-	assay at	Pseudomonas putida

General Information

General Information	Comment	Organism
metabolism	maleate isomerase catalyses the last step in nicotine degradation, the cis-trans isomerization of maleate to fumarate	Pseudomonas putida
additional information	residues C84 and C200 are essential for catalysis, active site structure, overview. The beta2-alpha2 loop and the beta6-alpha7 loop of Pp-Iso have a breathing motion revealed by the molecular dynamics simulation	Pseudomonas putida