Literature summary for 3.1.3.8 extracted from

Ranjan, B.; Singh, B.; Satyanarayana, T.
Characteristics of recombinant phytase (rSt-Phy) of the thermophilic mold Sporotrichum thermophile and its applicability in dephytinizing foods (2015), Appl. Biochem. Biotechnol., 177, 1753-1766 .

Activating Compound

Activating Compound	Comment	Organism
butanol	slight activation	Thermothelomyces thermophilus
isoamyl alcohol	slight activation	Thermothelomyces thermophilus
additional information	poor activation by hexane	Thermothelomyces thermophilus
Tween 20	34% activation at 0.5%	Thermothelomyces thermophilus
Tween 40	42% activation at 0.5%	Thermothelomyces thermophilus
Tween 80	67.5% activation at 0.5%	Thermothelomyces thermophilus

Application

Application	Comment	Organism
food industry	the recombinant enzyme rSt-Phy is useful in dephytinization of tandoori and naan (unleavened flat Indian breads), and bread, liberating soluble inorganic phosphate that mitigates anti-nutrient effects of phytic acid	Thermothelomyces thermophilus

Cloned(Commentary)

Cloned (Comment)	Organism
gene PHYA, construction of rSt-Phy gene using overlap extension PCR for removal of an intron, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21 (DE3), subcloning in Escherichia coli strain DH5alpha	Thermothelomyces thermophilus

Inhibitors

Inhibitors	Comment	Organism
2,3-Butanedione	-	Thermothelomyces thermophilus
Al3+	-	Thermothelomyces thermophilus
benzyl alcohol	slight inhibition	Thermothelomyces thermophilus
Ca2+	slight inhibition	Thermothelomyces thermophilus
Co2+	slight inhibition	Thermothelomyces thermophilus
Cu2+	-	Thermothelomyces thermophilus
dithiothreitol	slight inhibition	Thermothelomyces thermophilus
ethanol	slight inhibition	Thermothelomyces thermophilus
Fe2+	-	Thermothelomyces thermophilus
Fe3+	-	Thermothelomyces thermophilus
guanidinium hydrochloride	-	Thermothelomyces thermophilus
iodoacetate	slight inhibition	Thermothelomyces thermophilus
Mn2+	-	Thermothelomyces thermophilus
additional information	the enzyme is resistent against pepsin and trypsin, 95% of initial activity is retrieved after 1.0-h incubation in pepsin/trypsin. Poor inhibition by acetone, EDTA and 2-mercaptoethanol	Thermothelomyces thermophilus
N-bromosuccinimide	slight inhibition	Thermothelomyces thermophilus
N-ethylmaleimide	slight inhibition	Thermothelomyces thermophilus
Na+	slight inhibition	Thermothelomyces thermophilus
PMSF	-	Thermothelomyces thermophilus
potassium iodide	-	Thermothelomyces thermophilus
SDS	slight inhibition at 0.5%	Thermothelomyces thermophilus
Toluene	slight inhibition	Thermothelomyces thermophilus
Urea	-	Thermothelomyces thermophilus
Woodward's reagent K	-	Thermothelomyces thermophilus
Zn2+	-	Thermothelomyces thermophilus

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
additional information	-	additional information	thermodynamics of the recombinant enzyme	Thermothelomyces thermophilus
0.143	-	myo-inositol hexakisphosphate	recombinant enzyme, pH 5.0, 60°C	Thermothelomyces thermophilus
0.156	-	myo-inositol hexakisphosphate	native enzyme, pH 5.0, 60°C	Thermothelomyces thermophilus

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Ba2+	slight activation	Thermothelomyces thermophilus
Mg2+	slight activation	Thermothelomyces thermophilus

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
myo-inositol hexakisphosphate + H2O	Thermothelomyces thermophilus	-	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?
myo-inositol hexakisphosphate + H2O	Thermothelomyces thermophilus ATCC 42464	-	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?
myo-inositol hexakisphosphate + H2O	Thermothelomyces thermophilus BCRC 31852	-	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?
myo-inositol hexakisphosphate + H2O	Thermothelomyces thermophilus DSM 1799	-	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?

Organism

Organism	UniProt	Comment	Textmining
Thermothelomyces thermophilus	O00107	i.e. Sporotrichum thermophile or Thielavia heterothallica	-
Thermothelomyces thermophilus ATCC 42464	O00107	i.e. Sporotrichum thermophile or Thielavia heterothallica	-
Thermothelomyces thermophilus BCRC 31852	O00107	i.e. Sporotrichum thermophile or Thielavia heterothallica	-
Thermothelomyces thermophilus DSM 1799	O00107	i.e. Sporotrichum thermophile or Thielavia heterothallica	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant refolded His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and dialysis	Thermothelomyces thermophilus

Renatured (Commentary)

Renatured (Comment)	Organism
recombinant enzyme from Escherichia coli inclusion bodies is solubilized by 6 M urea and 5 mM 2-mercaptoethanol in 100 mM Tris-HCl, pH 7.0. The cell-free lysate is added to the equilibrated resins and is kept for 1 h on a rocker. The recombinant protein bound to the column is refolded on the column by passing five column volumes of buffer A (6 M urea and 5 mM 2-mercaptoethanol in 100 mM Tris-HCl, pH 7.0) and buffer B (20% v/v glycerol and 5 mM 2-mercaptoethanol in 100 mM Tris-HCl, pH 7.0) in a linear gradient of 6.0 to 0.0 M urea	Thermothelomyces thermophilus

Renatured (Comment)

Organism

recombinant enzyme from Escherichia coli inclusion bodies is solubilized by 6 M urea and 5 mM 2-mercaptoethanol in 100 mM Tris-HCl, pH 7.0. The cell-free lysate is added to the equilibrated resins and is kept for 1 h on a rocker. The recombinant protein bound to the column is refolded on the column by passing five column volumes of buffer A (6 M urea and 5 mM 2-mercaptoethanol in 100 mM Tris-HCl, pH 7.0) and buffer B (20% v/v glycerol and 5 mM 2-mercaptoethanol in 100 mM Tris-HCl, pH 7.0) in a linear gradient of 6.0 to 0.0 M urea

Thermothelomyces thermophilus

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg]	Specific Activity Maximum [µmol/min/mg]	Comment	Organism
52.4	-	purified native enzyme, pH 5.0, 60°C	Thermothelomyces thermophilus
8000	-	purified recombinant enzyme, pH 5.0, 60°C	Thermothelomyces thermophilus

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
myo-inositol hexakisphosphate + H2O	-	Thermothelomyces thermophilus	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?
myo-inositol hexakisphosphate + H2O	-	Thermothelomyces thermophilus ATCC 42464	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?
myo-inositol hexakisphosphate + H2O	-	Thermothelomyces thermophilus BCRC 31852	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?
myo-inositol hexakisphosphate + H2O	-	Thermothelomyces thermophilus DSM 1799	1D-myo-inositol 1,2,4,5,6-pentakisphosphate + phosphate	-	?

Subunits

Subunits	Comment	Organism
?	x * 53000, sequence calculation, x * 55000, recombinant His-tagged enzyme, SDS-PAGE, x * 90000, native enzyme, SDS-PAGE	Thermothelomyces thermophilus

Synonyms

Synonyms	Comment	Organism
3-phytase A	-	Thermothelomyces thermophilus
PhyA	-	Thermothelomyces thermophilus
St-Phy	-	Thermothelomyces thermophilus

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
60	-	recombinant and native enzymes	Thermothelomyces thermophilus

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
60	-	purified recombinant His-tagged enzyme, pH 5.0, has a half-life of 4 h. Purified native enzyme, pH 5.0, has a half-life of 16 h	Thermothelomyces thermophilus
80	-	purified recombinant His-tagged enzyme, pH 5.0, has a half-life of 38 min. Purified native enzyme, pH 5.0, has a half-life of 1.5 h	Thermothelomyces thermophilus

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
5100	-	myo-inositol hexakisphosphate	recombinant enzyme, pH 5.0, 60°C	Thermothelomyces thermophilus

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
5	-	recombinant and native enzymes	Thermothelomyces thermophilus

pI Value

Organism	Comment	pI Value Maximum	pI Value
Thermothelomyces thermophilus	sequence calculation	-	5.05

kcat/KM [mM/s]

kcat/KM Value [1/mMs^-1]	kcat/KM Value Maximum [1/mMs^-1]	Substrate	Comment	Organism	Structure
242.3	-	myo-inositol hexakisphosphate	native enzyme, pH 5.0, 60°C	Thermothelomyces thermophilus
37207.6	-	myo-inositol hexakisphosphate	recombinant enzyme, pH 5.0, 60°C	Thermothelomyces thermophilus