Literature summary for 4.1.1.47 extracted from

Kaplun, A.; Binshtein, E.; Vyazmensky, M.; Steinmatz, A.; Barak, Z.; Chipman, D.M.; Tittmann, K.; Shaanan, B.
Glyoxylate carboligase lacks the canonical active site glutamate of thiamine-dependent enzymes (2008), Nat. Chem. Biol., 4, 113-118.

Cloned(Commentary)

Cloned (Comment)	Organism
expression in Escherichia coli	Escherichia coli K-12

Crystallization (Commentary)

Crystallization (Comment)	Organism
glyoxylate carboligase crystallizes with 6 monomers (a tetramer and a dimer) in an asymmetric unit, vapor diffusion hanging-drop, 2-4 microlitre of protein solution (5-15 mg/ml, 100 micromolar ThDP, 10 micromolar FAD, 1mM MgCl2 and 10 mM quinone Q0) are mixed with equal volume of reservoir solution (0.5% PEG6000, 0.5M NaCl, 40 mM DTT), pH 8.00, temperature 294K, space group P41212, resolution 2.70 A	Escherichia coli K-12

Crystallization (Comment)

Organism

glyoxylate carboligase crystallizes with 6 monomers (a tetramer and a dimer) in an asymmetric unit, vapor diffusion hanging-drop, 2-4 microlitre of protein solution (5-15 mg/ml, 100 micromolar ThDP, 10 micromolar FAD, 1mM MgCl2 and 10 mM quinone Q0) are mixed with equal volume of reservoir solution (0.5% PEG6000, 0.5M NaCl, 40 mM DTT), pH 8.00, temperature 294K, space group P41212, resolution 2.70 A

Escherichia coli K-12

Protein Variants

Protein Variants	Comment	Organism
E52Q	site-directed mutagenesis	Escherichia coli K-12
V51D	site-directed mutagenesis	Escherichia coli K-12
V51E	site-directed mutagenesis	Escherichia coli K-12
V51S	site-directed mutagenesis	Escherichia coli K-12

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
0.2	-	glyoxylate	mutant V51D	Escherichia coli K-12
0.54	-	glyoxylate	mutant V51E	Escherichia coli K-12
0.9	-	glyoxylate	wild-type enzyme	Escherichia coli K-12
1.1	-	glyoxylate	mutant V51S	Escherichia coli K-12
1.2	-	glyoxylate	mutant E52Q	Escherichia coli K-12

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	-	Escherichia coli K-12

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
2 glyoxylate	Escherichia coli K-12	-	tartronate semialdehyde + CO2	-	?

Organism

Organism	UniProt	Comment	Textmining
Escherichia coli K-12	P0AEP7	-	-

Purification (Commentary)

Purification (Comment)	Organism
nickel affinity chromatography	Escherichia coli K-12

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
0.2	-	mutant V51D catalyzes the formation of the product nearly two orders of magnitude more slowly than the wild-type enzyme	Escherichia coli K-12
2.7	-	mutant V51E is about seven times slower than the wild-type enzyme	Escherichia coli K-12
17.1	-	mutant V51S is nearly as active as the wild-type enzyme	Escherichia coli K-12
17.5	-	wild-type enzyme	Escherichia coli K-12
18.3	-	mutant E52Q	Escherichia coli K-12

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
2 glyoxylate	-	Escherichia coli K-12	tartronate semialdehyde + CO2	-	?

Subunits

Subunits	Comment	Organism
homotetramer	the apparent functional unit is a dimer with a pair of identical thiamine diphosphate binding sites at the dimer interface	Escherichia coli K-12

Synonyms

Synonyms	Comment	Organism
GCL	-	Escherichia coli K-12
Glyoxylate carboligase	-	Escherichia coli K-12

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
0.25	-	glyoxylate	mutant V51D	Escherichia coli K-12
2.9	-	glyoxylate	mutant V51E	Escherichia coli K-12
18.5	-	glyoxylate	mutant V51S	Escherichia coli K-12
18.9	-	glyoxylate	wild-type enzyme	Escherichia coli K-12
19.7	-	glyoxylate	mutant E52Q	Escherichia coli K-12

Cofactor

Cofactor	Comment	Organism	Structure
FAD	-	Escherichia coli K-12
thiamine diphosphate	-	Escherichia coli K-12

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism
1.3	-	glyoxylate	mutant V51D	Escherichia coli K-12
5.3	-	glyoxylate	mutant V51E	Escherichia coli K-12
16.5	-	glyoxylate	mutant E52Q	Escherichia coli K-12
17.3	-	glyoxylate	mutant V51S	Escherichia coli K-12
21	-	glyoxylate	wild-type enzyme	Escherichia coli K-12