Literature summary for 1.13.11.8 extracted from

Barry, K.P.; Ngu, A.; Cohn, E.F.; Cote, J.M.; Burroughs, A.M.; Gerbino, J.P.; Taylor, E.A.
Exploring allosteric activation of LigAB from Sphingobium sp. strain SYK-6 through kinetics, mutagenesis and computational studies (2015), Arch. Biochem. Biophys., 567, 35-45 .

Search for more literature for 1.13.11.8

Activating Compound

Activating Compound	Comment	Organism	Structure
additional information	allosteric rate enhancement in the presence of non-substrate protocatechuate-like aldehydes such as vanillin	Sphingobium sp.
Vanillin	heteroallosteric activation of LigAB by vanillin at an allosteric binding site in LigAB, regulatory feed-forward activation mechanism, kinetics, overview. Vanillin is the pre-pre-substrate of LigAB. Enhancement by vanillin is specific to protocatechuate. Glu86b is directly involved in a hydrogen bond network through the carboxylate with Ndelta of His127b, believed to be the catalytic base that performs the initial substrate deprotonation. The His127b methylene also contributes a small fraction to the identified allosteric binding pocket	Sphingobium sp.

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of His6-tagged wild-type and mutant enzymes in Escherichia coli strain DH5alpha	Sphingobium sp.

Protein Variants

Protein Variants	Comment	Organism
A18W	site-directed mutagenesis, mutation in the betaa-subunit, residue Ala18b is located opening of the putative allosteric pocket, the mutant displays reduced activity and altered rate enhancement by vanillin compared to the wild-type enzyme	Sphingobium sp.
F103H	site-directed mutagenesis, mutation in the alpha-subunit, residue Phe103a is located in the putative allosteric pocket, the mutant displays reduced activity and altered rate enhancement by vanillin compared to the wild-type enzyme	Sphingobium sp.
F103L	site-directed mutagenesis, mutation in the alpha-subunit, residue Phe103a is located in the putative allosteric pocket, the mutant displays reduced activity and altered rate enhancement by vanillin compared to the wild-type enzyme	Sphingobium sp.
F103T	site-directed mutagenesis, mutation in the alpha-subunit, residue Phe103a is located in the putative allosteric pocket, the mutant displays reduced activity and altered rate enhancement by vanillin compared to the wild-type enzyme	Sphingobium sp.
F103V	site-directed mutagenesis, mutation in the alpha-subunit, residue Phe103a is located in the putative allosteric pocket, the mutant displays reduced activity and altered rate enhancement by vanillin compared to the wild-type enzyme	Sphingobium sp.

Inhibitors

Inhibitors	Comment	Organism	Structure
protocatechuate	substrate inhibition	Sphingobium sp.

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
additional information	-	additional information	Michaelis-Menten steady-state kinetics measuring O2 consumption	Sphingobium sp.
0.066	-	protocatechuate	pH 7.5, 25°C, recombinant mutant A18W	Sphingobium sp.
0.081	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme	Sphingobium sp.
0.108	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103H	Sphingobium sp.
0.13	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme, with vanillin	Sphingobium sp.
0.21	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103T	Sphingobium sp.
0.26	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103V	Sphingobium sp.
1.6	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103L	Sphingobium sp.

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
protocatechuate + O2	Sphingobium sp.	-	4-carboxy-2-hydroxymuconate semialdehyde	-	?

Organism

Organism	UniProt	Comment	Textmining
Sphingobium sp.	G2IQQ4 AND G2IQQ3	alpha- and beta-chain of LigAB; formerly Sphingomonas paucimobilis SYK-6	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His6-tagged wild-type and mutant enzymes anaerobically from Escherichia coli strain DH5alpha by nickel affinity chromatography, tag cleavage anaerobically by thrombin cleavage	Sphingobium sp.

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
additional information	LigAB has the broadest substrate utilization profile	Sphingobium sp.	?	-	?
protocatechuate + O2	-	Sphingobium sp.	4-carboxy-2-hydroxymuconate semialdehyde	-	?

Synonyms

Synonyms	Comment	Organism
LigAB	-	Sphingobium sp.

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Sphingobium sp.

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
33	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103L	Sphingobium sp.
89	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103V	Sphingobium sp.
110	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103H	Sphingobium sp.
112	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103T	Sphingobium sp.
122	-	protocatechuate	pH 7.5, 25°C, recombinant mutant A18W	Sphingobium sp.
178	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme	Sphingobium sp.
230	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme, with vanillin	Sphingobium sp.

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Sphingobium sp.

Ki Value [mM]

Ki Value [mM]	Ki Value maximum [mM]	Inhibitor	Comment	Organism	Structure
17	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme, with vanillin	Sphingobium sp.
18	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme	Sphingobium sp.

General Information

General Information	Comment	Organism
evolution	protocatechuate 4,5-dioxygenase (LigAB) from Sphingobium sp. strain SYK-6 is the defining member of the type II extradiol dioxygenase superfamily, a.k.a. PCA dioxygenase superfamily or PCADSF, computational docking and sequence comparisons of the enzyme with PCA dioxygenase superfamily LigAB enzymes	Sphingobium sp.
physiological function	the enzyme plays a key aromatic ring-opening role in the metabolism of several lignin derived aromatic compounds	Sphingobium sp.

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
21	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103L	Sphingobium sp.
340	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103V	Sphingobium sp.
530	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103T	Sphingobium sp.
1100	-	protocatechuate	pH 7.5, 25°C, recombinant mutant F103H	Sphingobium sp.
1800	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme, with vanillin	Sphingobium sp.
1900	-	protocatechuate	pH 7.5, 25°C, recombinant mutant A18W	Sphingobium sp.
2200	-	protocatechuate	pH 7.5, 25°C, recombinant wild-type enzyme	Sphingobium sp.

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Release 2023.1 (January 2023)