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Literature summary extracted from

  • Lin, H.Y.; Chuang, H.H.; Lin, F.P.
    Biochemical characterization of engineered amylopullulanase from Thermoanaerobacter ethanolicus 39E-implicating the non-necessity of its 100 C-terminal amino acid residues (2008), Extremophiles, 12, 641-650.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.2.1.135 Escherichia coli NovaBlue and Rosetta (DE3) pLysS, cloning and expression host, respectively, with plasmid pET-20b(+), in LB broth, 37°C Thermoanaerobacter ethanolicus

Protein Variants

EC Number Protein Variants Comment Organism
3.2.1.135 additional information R855 C-terminal truncated (100 amino acids) mutant R855 of full length TetApuM955, produced with trypsin-like proteolytic cleavage reaction Thermoanaerobacter ethanolicus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.2.1.135 additional information
-
pullulan 11.6 mg/ml, TetApuM955 Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
pullulan 16.0 mg/ml, TetApuR855 Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
starch 5.2 mg/ml, TetApuM955 Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
starch 6.0 mg/ml, TetApuR855 Thermoanaerobacter ethanolicus

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
3.2.1.135 85000
-
theoretically calculated mass of truncated TetApuM855, corroborated by SDS-PAGE Thermoanaerobacter ethanolicus
3.2.1.135 107800
-
calculated mass of matured full length enzmye TetAbuM955, corroborated by SDS-PAGE, 13 N-terminal amino acids (signal peptide) cleaved off proteolytically, DNA fragment 2882 bp Thermoanaerobacter ethanolicus
3.2.1.135 107818
-
x * 107818, SDS-PAGE Thermoanaerobacter ethanolicus

Organism

EC Number Organism UniProt Comment Textmining
3.2.1.135 Thermoanaerobacter ethanolicus
-
TetApuM955 gene of 2.9 kb
-
3.2.1.135 Thermoanaerobacter ethanolicus 39E
-
TetApuM955 gene of 2.9 kb
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.2.1.135 BD TALON spin column for 6x His-tag protein affinity purification, simultaneous SDS-PAGE zymogram with 1% soluble starch Thermoanaerobacter ethanolicus

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.2.1.135 additional information
-
117400000000 micromol/min/mol enzyme, TetApuR855, soluble starch Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
11750000000 micromol/min/mol enzyme, TetApuR855, raw starch Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
11860000000 micromol/min/mol enzyme, TetApuM955, raw starch Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
119400000000 micromol/min/mol enzyme, TetApuM955, soluble starch Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
234200000000 micromol/min/mol enzyme, TetApuM955, pullulan Thermoanaerobacter ethanolicus
3.2.1.135 additional information
-
243000000000 micromol/min/mol enzyme, TetApuR855, pullulan Thermoanaerobacter ethanolicus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.2.1.135 glycogen + H2O 50 mM sodium acetate buffer, pH 6.0, 5 mM CaCl2, 80°C Thermoanaerobacter ethanolicus maltooligosaccharides
-
?
3.2.1.135 glycogen + H2O 50 mM sodium acetate buffer, pH 6.0, 5 mM CaCl2, 80°C Thermoanaerobacter ethanolicus 39E maltooligosaccharides
-
?
3.2.1.135 pullulan + H2O 50 mM sodium acetate buffer, pH 6.0, 5 mM CaCl2, 75°C Thermoanaerobacter ethanolicus maltooligosaccharides
-
?
3.2.1.135 pullulan + H2O 50 mM sodium acetate buffer, pH 6.0, 5 mM CaCl2, 75°C Thermoanaerobacter ethanolicus 39E maltooligosaccharides
-
?
3.2.1.135 soluble and raw starch + H2O 50 mM sodium acetate buffer, pH 6.0, 5 mM CaCl2, 80°C Thermoanaerobacter ethanolicus maltose + maltotriose + maltotetraose + maltopentaose + maltohexaose + maltoheptaose + ?
-
?
3.2.1.135 soluble and raw starch + H2O 50 mM sodium acetate buffer, pH 6.0, 5 mM CaCl2, 80°C Thermoanaerobacter ethanolicus 39E maltose + maltotriose + maltotetraose + maltopentaose + maltohexaose + maltoheptaose + ?
-
?
3.2.1.135 starch + H2O
-
Thermoanaerobacter ethanolicus maltose + ?
-
?
3.2.1.135 starch + H2O
-
Thermoanaerobacter ethanolicus 39E maltose + ?
-
?

Subunits

EC Number Subunits Comment Organism
3.2.1.135 ? x * 85000, SDS-PAGE Thermoanaerobacter ethanolicus
3.2.1.135 ? x * 107818, SDS-PAGE Thermoanaerobacter ethanolicus

Synonyms

EC Number Synonyms Comment Organism
3.2.1.135 amylopullulanase
-
Thermoanaerobacter ethanolicus
3.2.1.135 TetApuM955 full length enzyme Thermoanaerobacter ethanolicus
3.2.1.135 TetApuR855 truncated enzyme Thermoanaerobacter ethanolicus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.2.1.135 75
-
75-80°C optimal for full length and truncated enzyme Thermoanaerobacter ethanolicus

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
3.2.1.135 80
-
no temperature sensitivity difference between truncated and full length enzyme below and at 80°C Thermoanaerobacter ethanolicus
3.2.1.135 88
-
onset of denaturation of full length TetApuM955 Thermoanaerobacter ethanolicus
3.2.1.135 90
-
onset of denaturation of truncated TetApuR885, it is more thermostable than full length TetApuM955 above 90°C Thermoanaerobacter ethanolicus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.2.1.135 298.3
-
starch TetApuM955 Thermoanaerobacter ethanolicus
3.2.1.135 313.3
-
starch TetApuR855 Thermoanaerobacter ethanolicus
3.2.1.135 828.3
-
pullulan TetApuM955 Thermoanaerobacter ethanolicus
3.2.1.135 1010
-
pullulan TetApuR855 Thermoanaerobacter ethanolicus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.2.1.135 6
-
full length and truncated enzyme Thermoanaerobacter ethanolicus