Literature summary for 5.1.1.1 extracted from

Tassoni, R.; van der Aart, L.T.; Ubbink, M.; van Wezel, G.P.; Pannu, N.S.
Structural and functional characterization of the alanine racemase from Streptomyces coelicolor A3(2) (2017), Biochem. Biophys. Res. Commun., 483, 122-128 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene alr, sequence comparisons, recombinant expression of N-terminally His6-tagged enzyme in Escherichia coli strain BL21 Star (DE3)pLysS	Streptomyces coelicolor

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant N-terminally His-tagged enzyme free, or in complex with inhibitors D-cycloserine and propionate, sitting drop vapor diffusion, mixing of 0.001 ml of 20 mg/ml protein solution with 0.001 ml of crystallization solution containing 0.1 M Bis-Tris propane, pH 8.5, 0.2 M NaBr, 20% w/v PEG 3350, 38 days, X-ray diffraction structure determination and analysis at 2.8 A for the free enzyme, and at 1.51-1.64 A resolution for the enzyme complexes, model building	Streptomyces coelicolor

Crystallization (Comment)

Organism

purified recombinant N-terminally His-tagged enzyme free, or in complex with inhibitors D-cycloserine and propionate, sitting drop vapor diffusion, mixing of 0.001 ml of 20 mg/ml protein solution with 0.001 ml of crystallization solution containing 0.1 M Bis-Tris propane, pH 8.5, 0.2 M NaBr, 20% w/v PEG 3350, 38 days, X-ray diffraction structure determination and analysis at 2.8 A for the free enzyme, and at 1.51-1.64 A resolution for the enzyme complexes, model building

Streptomyces coelicolor

Inhibitors

Inhibitors	Comment	Organism	Structure
D-cycloserine	structural features such as the hinge angle or the surface area between the monomers do not contribute to D-cycloserine resistance, binding structure analysis, overview	Streptomyces coelicolor
propionate	binding structure analysis, overview	Streptomyces coelicolor

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
83000	-	gel filtration, recombinant enzyme	Streptomyces coelicolor

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
L-alanine	Streptomyces coelicolor	-	D-alanine	-	r
L-alanine	Streptomyces coelicolor ATCC BAA-471 / A3(2) / M145	-	D-alanine	-	r

Organism

Organism	UniProt	Comment	Textmining
Streptomyces coelicolor	O86786	-	-
Streptomyces coelicolor ATCC BAA-471 / A3(2) / M145	O86786	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant N-terminally His6-tagged enzyme from Escherichia coli strain BL21 Star (DE3)pLysS by nickel affinity chromatography	Streptomyces coelicolor

Source Tissue

Source Tissue	Comment	Organism	Textmining
mycelium	-	Streptomyces coelicolor	-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
66	-	racemization of L- to D-Ala, purified enzyme, pH 7.0, 22°C	Streptomyces coelicolor
104	-	racemization of D- to L-Ala, purified enzyme, pH 7.0, 22°C	Streptomyces coelicolor

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
L-alanine	-	Streptomyces coelicolor	D-alanine	-	r
L-alanine	-	Streptomyces coelicolor ATCC BAA-471 / A3(2) / M145	D-alanine	-	r

Subunits

Subunits	Comment	Organism
homodimer	2 * 43400, recombinant enzyme, SDS-PAGE	Streptomyces coelicolor
More	enzyme Alr is a homodimer with residues from both monomers contributing to the active site. There are two active sites per dimer, which are located at the interface between each alpha/beta-barrel of one subunit and the C-terminal domain of the other. The catalytic core consists of the pyridoxal 5'-phosphate cofactor, a Lys, and a Tyr, which is contributed by the other subunit. The cofactor is bound through an internal aldimine bond to the amino group of Lys46, located at the C-terminal side of the first beta-strand of the alpha/beta-barrel. The side chain of the catalytic Lys46 points out of the alpha/beta-barrel, towards the C-terminal domain of the interacting subunit, and in particular, towards Tyr283'. The phosphate group of the pyridoxal-5'-phosphate is stabilized by hydrogen bonds with the side chains of Tyr50, Ser222 and Tyr374, and with the backbone of Gly239, Ser222, and Ile240	Streptomyces coelicolor

Synonyms

Synonyms	Comment	Organism
ALR	-	Streptomyces coelicolor

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
22	-	assay at room temperature	Streptomyces coelicolor

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
95	-	purified recombinant enzyme, pH 7.0, 3 min, inactivation	Streptomyces coelicolor

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7	-	assay at	Streptomyces coelicolor

Cofactor

Cofactor	Comment	Organism	Structure
pyridoxal 5'-phosphate	PLP, dependent on, binding structure analysis: the phosphate group of the pyridoxal 5'-phosphate is stabilized by hydrogen bonds with the side chains of Tyr50, Ser222 and Tyr374, and with the backbone of Gly239, Ser222, and Ile240. The pyridine ring of the PLP is stabilized by a hydrogen bond between the N-1 of the cofactor and Nepsilon of Arg237. The C2A of the PLP also interacts with oxygen Q1 of the carboxylated Lys141. All residues stabilizing the PLP cofactor (Tyr50, Ser222, Gly239, Ile240, Arg237, Tyr374) are conserved among Alr proteins. But the AlrSco lacks one important hydrogen bond between Arg148 and the phenolic oxygen of the PLP molecule	Streptomyces coelicolor

General Information

General Information	Comment	Organism
evolution	the enzyme belongs to the Fold Type III of pyridoxal 5'-phosphate-dependent enzymes	Streptomyces coelicolor
additional information	structure-function realtionship analysis, overview	Streptomyces coelicolor
physiological function	the conversion of L-alanine (L-Ala) into D-alanine (D-Ala) in bacteria is performed by pyridoxal 5'-phosphate-dependent enzymes, alanine racemases. D-Ala is an essential component of the bacterial peptidoglycan and hence required for survival	Streptomyces coelicolor