Literature summary for 3.1.1.81 extracted from

Hiblot, J.; Bzdrenga, J.; Champion, C.; Chabriere, E.; Elia, M.
Crystal structure of VmoLac, a tentative quorum quenching lactonase from the extremophilic crenarchaeon Vulcanisaeta moutnovskia (2015), Sci. Rep., 5, 8372 .

Cloned(Commentary)

Cloned (Comment)	Organism
gene VMUT_2255, phylogenetic analysis and sequence comparisons, recombinant enzyme expression in Escherichia coli strain BL21(DE3)-pGro7/GroEL	Vulcanisaeta moutnovskia

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant enzyme, with or without substrate, sitting drop vapor diffusion method, mixing of 20 mg/ml protein with different crystallization solutions, i.e. 400 mM ammonium dihydrogen phosphate plus G9 (100 mM sodium citrate, pH 5.6, 10 mM ferric chloride, and 10% v/v jeffamine M-600), C4 (100 mM sodium HEPES, pH 7.5, 800 mM sodium, and potassium dihydrogen phosphate) or E7 (100 mM Tris, pH 8.5, 1.5 M ammonium sulfate, and 12% v/v glycerol) in a 2:1 and 1:1 protein to reservoir ratio. The crystals from the G9 and E7 conditions have the same P64 space group and the ones issued from the C4 condition have an alternative P622 group space. The crystals from E7 are soaked into the well solution supplemented by 2 mM of substrate N-oxodecanoyl-DL-homoserine lactone for 1 min prior to flash cooling, crystal growth at °C, 2 weeks, X-ray diffraction structure determination and analysis at 1.8-2.35 A resolution, molecular replacement using the SsoPox structure as a starting template (PDB ID 2vc5), modelling	Vulcanisaeta moutnovskia

Crystallization (Comment)

Organism

purified recombinant enzyme, with or without substrate, sitting drop vapor diffusion method, mixing of 20 mg/ml protein with different crystallization solutions, i.e. 400 mM ammonium dihydrogen phosphate plus G9 (100 mM sodium citrate, pH 5.6, 10 mM ferric chloride, and 10% v/v jeffamine M-600), C4 (100 mM sodium HEPES, pH 7.5, 800 mM sodium, and potassium dihydrogen phosphate) or E7 (100 mM Tris, pH 8.5, 1.5 M ammonium sulfate, and 12% v/v glycerol) in a 2:1 and 1:1 protein to reservoir ratio. The crystals from the G9 and E7 conditions have the same P64 space group and the ones issued from the C4 condition have an alternative P622 group space. The crystals from E7 are soaked into the well solution supplemented by 2 mM of substrate N-oxodecanoyl-DL-homoserine lactone for 1 min prior to flash cooling, crystal growth at °C, 2 weeks, X-ray diffraction structure determination and analysis at 1.8-2.35 A resolution, molecular replacement using the SsoPox structure as a starting template (PDB ID 2vc5), modelling

Vulcanisaeta moutnovskia

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
additional information	-	additional information	gamma- and delta-dodecanoic lactone substrates have an allosteric profile of hydrolysis	Vulcanisaeta moutnovskia
0.0021	-	dodecanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.047	-	nonanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.105	-	undecanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.12	-	undecanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.186	-	N-3-oxo-octanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.231	-	N-3-oxo-decanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.262	-	N-octanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.55	-	(R)-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.75	-	(S)-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.839	-	dodecanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.872	-	heptanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
1.22	-	nonanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
3.709	-	rac-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Co2+	required, VmoLac is a bi-cobalt metalloenzyme. The two metal cations are coordinated by four histidines, His23, His25, His171, and His200, an aspartic acid Asp257, and a carboxylated lysine residue Lys238	Vulcanisaeta moutnovskia
additional information	the enzyme contains a bi-metallic active site that is coordinated by four histidines, an aspartic acid and a carboxylated lysine. The bi-metallic center acts as a Lewis acid and activates a bridging, putatively catalytic, water molecule into a hydroxide anion, which subsequently serves as a nucleophile for the organophosphorous compounds or lactone hydrolysis	Vulcanisaeta moutnovskia

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
an N-acyl-L-homoserine lactone + H2O	Vulcanisaeta moutnovskia	-	an N-acyl-L-homoserine	-	?
homo-L-cysteine thiolactone + H2O	Vulcanisaeta moutnovskia	-	homo-L-cysteine	-	?

Organism

Organism	UniProt	Comment	Textmining
Vulcanisaeta moutnovskia	F0QXN6	a hyperthermophilic crenarchaeon	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
(R)-gamma-caprolactone + H2O	-	Vulcanisaeta moutnovskia	4-hydroxycaproate	-	?
(S)-gamma-caprolactone + H2O	-	Vulcanisaeta moutnovskia	caproate	-	?
an N-acyl-L-homoserine lactone + H2O	-	Vulcanisaeta moutnovskia	an N-acyl-L-homoserine	-	?
dodecanoic-delta-lactone + H2O	-	Vulcanisaeta moutnovskia	5-hydroxydodecanoate	-	?
dodecanoic-gamma-lactone + H2O	-	Vulcanisaeta moutnovskia	dodecanoate	-	?
epsilon-caprolactone + H2O	-	Vulcanisaeta moutnovskia	6-hydroxycaproate	-	?
gamma-heptanolide + H2O	-	Vulcanisaeta moutnovskia	?	-	?
heptanoic-gamma-lactone + H2O	-	Vulcanisaeta moutnovskia	4-hydroxyheptanoate	-	?
homo-L-cysteine thiolactone + H2O	-	Vulcanisaeta moutnovskia	homo-L-cysteine	-	?
additional information	the enzyme is also active on diethyl-paraoxon, dimethyl-paraoxon, diethyl-parathion, dimethyl-parathion, and malathion. The lactone ring hydrolysis is followed by acidification of the medium for which Cresol purple serves as pH indicator. No or poor activity is detected against phenylacetate, dihydrocoumarin, 4-nitrophenyl acetate, 4-nitrophenyl decanoate, and AHLs with short aliphatic substituents (4 to 6 carbon atoms), e.g. N-hexanoyl-DL-homoserine lactone, N-butanoyl-DL-homoserine lactone, and N-3-oxo-hexanoyl-DL-homoserine lactone. AHLs with longer substituents are better substrates for VmoLac, including C8 and C10-AHLs. Oxo-lactones are the best substrates for VmoLac. gamma-caprolactone and gamma-heptalactone are degraded with approximately the same catalytic efficiencies, and oxo-lactones with long aliphatic chains such as nonanoic-gamma-lactone and undecanoic-delta-lactone, are hydrolyzed with high catalytic efficiencies	Vulcanisaeta moutnovskia	?	-	?
N-3-oxo-decanoyl-DL-homoserine lactone + H2O	-	Vulcanisaeta moutnovskia	N-3-oxo-decanoyl-DL-homoserine	-	?
N-3-oxo-octanoyl-DL-homoserine lactone + H2O	-	Vulcanisaeta moutnovskia	N-3-oxo-octanoyl-DL-homoserine	-	?
N-octanoyl-DL-homoserine lactone + H2O	-	Vulcanisaeta moutnovskia	N-octanoyl-DL-homoserine	-	?
nonanoic-delta-lactone + H2O	-	Vulcanisaeta moutnovskia	5-hydroxynonanoate	-	?
nonanoic-delta-lactone + H2O	-	Vulcanisaeta moutnovskia	5-hydroxynonanoic acid	-	?
nonanoic-gamma-lactone + H2O	best substrate	Vulcanisaeta moutnovskia	4-hydroxynonanoate	-	?
nonanoic-gamma-lactone + H2O	best substrate	Vulcanisaeta moutnovskia	4-hydroxynonanoic acid	-	?
rac-gamma-caprolactone + H2O	-	Vulcanisaeta moutnovskia	4-hydroxycaproate	-	?
undecanoic-delta-lactone + H2O	-	Vulcanisaeta moutnovskia	5-hydroxyundecanoate	-	?
undecanoic-gamma-lactone + H2O	-	Vulcanisaeta moutnovskia	4-hydroxyundecanoate	-	?

Subunits

Subunits	Comment	Organism
More	VmoLac is roughly globular and exhibits a (beta/alpha)8 barrel topology	Vulcanisaeta moutnovskia

Synonyms

Synonyms	Comment	Organism
More	cf. EC 3.1.8.1	Vulcanisaeta moutnovskia
quorum quenching lactonase	-	Vulcanisaeta moutnovskia
VmoLac	-	Vulcanisaeta moutnovskia
VMUT_2255	-	Vulcanisaeta moutnovskia

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
40	-	assay at	Vulcanisaeta moutnovskia

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
0.16	-	dodecanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.35	-	N-3-oxo-octanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.47	-	N-3-oxo-decanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
0.59	-	N-octanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
1.89	-	(S)-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
3.04	-	(R)-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
8.86	-	undecanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
27.25	-	heptanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
28.08	-	dodecanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
44.49	-	nonanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
60.78	-	undecanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
88.91	-	nonanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
112.3	-	rac-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8.3	-	assay at	Vulcanisaeta moutnovskia

General Information

General Information	Comment	Organism
evolution	the enzyme belongs to the phosphotriesterase-like lactonases (PLLs) family of enzyme. Phosphotriesterase-like lactonases (PLLs) compose a class of lactonases that has long been mistaken for the organophosphorus-degrading enzymes phosphotriesterases (PTEs, EC 3.1.8.1). Several PLLs have initially been isolated and characterized by virtue of their organophosphorous insecticide degrading abilities. PLLs belong to the amidohydrolase superfamily and exhibit a (beta/alpha)8 topology containing a bi-metallic active site that is coordinated by four histidines, an aspartic acid and a carboxylated lysine. PTEs and PLLs mainly differ by the size and conformations of the active site loops 7 and 8 that mediate substrate specificity	Vulcanisaeta moutnovskia
additional information	the active site loops 7 and 8 mediate the substrate specificity. VmoLac possesses a bi-cobalt active site. Active site structure and structure comparisons, overview	Vulcanisaeta moutnovskia

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism
1.88	-	N-3-oxo-octanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
2.04	-	N-3-oxo-decanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
2.25	-	N-octanoyl-DL-homoserine lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
2.52	-	(S)-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
5.53	-	(R)-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
30.28	-	rac-gamma-caprolactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
31.25	-	heptanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
33.47	-	dodecanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
72.88	-	nonanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
73.83	-	undecanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
76.19	-	dodecanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
578.86	-	undecanoic-delta-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia
946.6	-	nonanoic-gamma-lactone	pH 8.0, 25°C, recombinant enzyme	Vulcanisaeta moutnovskia