Literature summary for 4.4.1.13 extracted from

Ferla, M.P.; Brewster, J.L.; Hall, K.R.; Evans, G.B.; Patrick, W.M.
Primordial-like enzymes from bacteria with reduced genomes (2017), Mol. Microbiol., 105, 508-524 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene metC, phylogenetic analysis, the recombinant His6-tagged enzyme TmCBL is overexpressed in a metC-deficient DELTAmetC Escherichia coli mutant strain and can complement it. The enzyme rescues the alanine racemase knockout, Escherichia coli strain MB2795 (DELTAalrDELTAdadX), as quickly as expressing Escherichia coli ALR itself. Recombinant production of His6-tagged enzyme in a soluble form	Thermotoga maritima
gene metC, phylogenetic analysis, the recombinant His6-tagged enzyme wMelCBL is overexpressed in a metC-deficient DELTAmetC Escherichia coli mutant strain and can complement it. The enzyme rescues the alanine racemase knockout, Escherichia coli strain MB2795 (DELTAalrDELTAdadX), as quickly as expressing Escherichia coli ALR itself. Recombinant production of His6-tagged enzyme in a soluble form	Wolbachia pipientis
gene metC, phylogenetic analysis, the recombinant MBP-tagged enzyme PuCBL is overexpressed in a metC-deficient DELTAmetC Escherichia coli mutant strain and can complement it partially. The enzyme rescues the alanine racemase knockout, Escherichia coli strain MB2795 (DELTAalrDELTAdadX), as quickly as expressing Escherichia coli ALR itself. Attempts to optimize recombinant production of the Pelagibacter ubique enzyme (PuCBL) in a soluble form are unsuccessful	Candidatus Pelagibacter ubique
gene metC, phylogenetic analysis. The recombinant enzyme EcCBL is overexpressed in a metC-deficient DELTAmetC Escherichia coli mutant strain and can complement it partially	Escherichia coli

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
additional information	-	additional information	Michaelis-Menten steady-state kinetics	Candidatus Pelagibacter ubique
additional information	-	additional information	Michaelis-Menten steady-state kinetics	Wolbachia pipientis
additional information	-	additional information	Michaelis-Menten steady-state kinetics	Thermotoga maritima
0.02	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C	Wolbachia pipientis
0.4	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C, Tris buffer	Thermotoga maritima
0.7	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C, bicine buffer	Thermotoga maritima
1.3	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 70°C, bicine buffer	Thermotoga maritima
2.1	-	cystathionine	recombinant MBP-tagged enzyme, pH 8.0, 37°C	Candidatus Pelagibacter ubique

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
cystathionine + H2O	Escherichia coli	-	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	Candidatus Pelagibacter ubique	-	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	Wolbachia pipientis	-	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	Thermotoga maritima	-	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	Candidatus Pelagibacter ubique HTCC1062	-	L-homocysteine + pyruvate + NH3	-	?
additional information	Wolbachia pipientis	the enzyme is a physiologically bifunctional alanine/glutamate racemase (EC 5.1.1.1/EC 5.1.1.3), it is not highly active, but it is clearly sufficient. The metC encoded L-alanine/L-glutamate racemase is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	?	-	-
additional information	Thermotoga maritima	the enzyme is a physiologically bifunctional alanine/glutamate racemase (EC 5.1.1.1/EC 5.1.1.3), it is not highly active, but it is clearly sufficient. The metC encoded L-alanine/L-glutamate racemase is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	?	-	-
additional information	Wolbachia pipientis wMel	the enzyme is a physiologically bifunctional alanine/glutamate racemase (EC 5.1.1.1/EC 5.1.1.3), it is not highly active, but it is clearly sufficient. The metC encoded L-alanine/L-glutamate racemase is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	?	-	-

Organism

Organism	UniProt	Comment	Textmining
Candidatus Pelagibacter ubique	Q4FME3	-	-
Candidatus Pelagibacter ubique HTCC1062	Q4FME3	-	-
Escherichia coli	P06721	-	-
Thermotoga maritima	Q9X0Z7	-	-
Thermotoga maritima ATCC 43589	Q9X0Z7	-	-
Thermotoga maritima DSM 3109	Q9X0Z7	-	-
Thermotoga maritima JCM 10099	Q9X0Z7	-	-
Wolbachia pipientis	Q73GL9	endosymbiont of Drosophila melanogaster	-
Wolbachia pipientis wMel	Q73GL9	endosymbiont of Drosophila melanogaster	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
cystathionine + H2O	-	Escherichia coli	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	-	Candidatus Pelagibacter ubique	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	-	Wolbachia pipientis	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	-	Thermotoga maritima	L-homocysteine + pyruvate + NH3	-	?
cystathionine + H2O	-	Candidatus Pelagibacter ubique HTCC1062	L-homocysteine + pyruvate + NH3	-	?
additional information	the enzyme is a physiologically bifunctional alanine/glutamate racemase (EC 5.1.1.1/EC 5.1.1.3), it is not highly active, but it is clearly sufficient. The metC encoded L-alanine/L-glutamate racemase is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	Wolbachia pipientis	?	-	-
additional information	the enzyme is a physiologically bifunctional alanine/glutamate racemase (EC 5.1.1.1/EC 5.1.1.3), it is not highly active, but it is clearly sufficient. The metC encoded L-alanine/L-glutamate racemase is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	Thermotoga maritima	?	-	-
additional information	no activity with O-acetylhomoserine or O-succinylhomoserine plus L-cysteine, the cystathionine gamma-synthase (metB, EC 2.5.1.48) activity of the enzyme is negligible	Thermotoga maritima	?	-	-
additional information	the Escherichia coli cystathionine beta-lyase (CBL) also shows a promiscuous alanine racemase (ALR) activity (cf. EC 5.1.1.1). ALR catalyzes the interconversion of L-alanine and D-alanine	Escherichia coli	?	-	-
additional information	the GLR (EC 5.1.1.3) activity of wMelCBL is very weak: approximately 3fold less efficient than cystathionine beta-elimination (EC 4.4.1.13) and 28fold less efficient than alanine racemization (EC 5.1.1.1)	Wolbachia pipientis	?	-	-
additional information	the Pelagibacter ubique cystathionine beta-lyase (CBL) also shows a promiscuous alanine racemase (ALR) activity (cf. EC 5.1.1.1). ALR catalyzes the interconversion of L-alanine and D-alanine. The ALR activity of MBP-PuCBL is 40fold lower than its CBL activity	Candidatus Pelagibacter ubique	?	-	-
additional information	the enzyme is a physiologically bifunctional alanine/glutamate racemase (EC 5.1.1.1/EC 5.1.1.3), it is not highly active, but it is clearly sufficient. The metC encoded L-alanine/L-glutamate racemase is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	Wolbachia pipientis wMel	?	-	-
additional information	the Pelagibacter ubique cystathionine beta-lyase (CBL) also shows a promiscuous alanine racemase (ALR) activity (cf. EC 5.1.1.1). ALR catalyzes the interconversion of L-alanine and D-alanine. The ALR activity of MBP-PuCBL is 40fold lower than its CBL activity	Candidatus Pelagibacter ubique HTCC1062	?	-	-

Subunits

Subunits	Comment	Organism
dimer	the Escherichia coli ALR dimer (PDB entry 2RJG) adopts a fold type III	Escherichia coli
tetramer	the Escherichia coli CBL tetramer (PDB entry 1CL1) adopts a fold type I	Escherichia coli

Synonyms

Synonyms	Comment	Organism
AAR	-	Wolbachia pipientis
AAR	-	Thermotoga maritima
CBL	-	Escherichia coli
CBL	-	Candidatus Pelagibacter ubique
CBL	-	Wolbachia pipientis
CBL	-	Thermotoga maritima
CBL/ALR	-	Escherichia coli
CBL/ALR	-	Candidatus Pelagibacter ubique
CBL/ALR	-	Wolbachia pipientis
CBL/ALR	-	Thermotoga maritima
cystathionine beta-lyase	-	Escherichia coli
cystathionine beta-lyase	-	Candidatus Pelagibacter ubique
cystathionine beta-lyase	-	Wolbachia pipientis
cystathionine beta-lyase	-	Thermotoga maritima
EcCBL	-	Escherichia coli
MetC	-	Escherichia coli
MetC	-	Candidatus Pelagibacter ubique
MetC	-	Wolbachia pipientis
MetC	-	Thermotoga maritima
More	see also EC 5.1.1.1	Escherichia coli
More	see also EC 5.1.1.1	Candidatus Pelagibacter ubique
More	see also EC 5.1.1.1 and EC 5.1.1.3	Wolbachia pipientis
More	see also EC 5.1.1.1 and EC 5.1.1.3	Thermotoga maritima
TmCBL	-	Thermotoga maritima
wMelCBL	-	Wolbachia pipientis

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Candidatus Pelagibacter ubique
37	-	assay at	Wolbachia pipientis
70	-	-	Thermotoga maritima

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
0.0011	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C	Wolbachia pipientis
1	-	cystathionine	recombinant MBP-tagged enzyme, pH 8.0, 37°C	Candidatus Pelagibacter ubique
2.3	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C, Tris buffer	Thermotoga maritima
3.3	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C, bicine buffer	Thermotoga maritima
9.2	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 70°C, bicine buffer	Thermotoga maritima

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8	-	assay at	Candidatus Pelagibacter ubique
8	-	assay at	Wolbachia pipientis
8	-	assay at	Thermotoga maritima

Cofactor

Cofactor	Comment	Organism
pyridoxal 5'-phosphate	-	Escherichia coli
pyridoxal 5'-phosphate	-	Candidatus Pelagibacter ubique
pyridoxal 5'-phosphate	-	Wolbachia pipientis
pyridoxal 5'-phosphate	-	Thermotoga maritima

General Information

General Information	Comment	Organism
evolution	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria. Primordial-like enzymes may be an essential part of the adaptive strategy associated with streamlining. Evolutionary history of CBL	Escherichia coli
evolution	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria. Primordial-like enzymes may be an essential part of the adaptive strategy associated with streamlining. Evolutionary history of CBL	Candidatus Pelagibacter ubique
evolution	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria. Primordial-like enzymes may be an essential part of the adaptive strategy associated with streamlining. Evolutionary history of CBL	Wolbachia pipientis
evolution	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria. Primordial-like enzymes may be an essential part of the adaptive strategy associated with streamlining. Evolutionary history of CBL	Thermotoga maritima
metabolism	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	Escherichia coli
metabolism	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	Candidatus Pelagibacter ubique
metabolism	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	Wolbachia pipientis
metabolism	several bacteria with reduced genomes lack alr, encoding alanine racemase (EC 5.1.1.1), but contain metC encoding cystathionine beta-lyase (CBL), which, in these organisms, is a multifunctional CBL/ALR. CBL activity is no longer required in these bacteria	Thermotoga maritima
additional information	comparisons of structure and function of CBL and ALR, overview	Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism
0.055	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C	Wolbachia pipientis
0.476	-	cystathionine	recombinant MBP-tagged enzyme, pH 8.0, 37°C	Candidatus Pelagibacter ubique
4.71	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C, bicine buffer	Thermotoga maritima
5.75	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 37°C, Tris buffer	Thermotoga maritima
7.08	-	cystathionine	recombinant His-tagged enzyme, pH 8.0, 70°C, bicine buffer	Thermotoga maritima