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Literature summary extracted from
- Crystal structure of the polyextremophilic alpha-amylase AmyB from Halothermothrix orenii: details of a productive enzyme-substrate complex and an N-domain with a role in binding raw starch (2008), J. Mol. Biol., 378, 852-870.
Cloned(Commentary)
| EC Number | Cloned (Comment) | Organism |
|---|---|---|
| 3.2.1.1 | expressed ion Escherichia coli BL21(DE3) | Halothermothrix orenii |
Crystallization (Commentary)
| EC Number | Crystallization (Comment) | Organism |
|---|---|---|
| 3.2.1.1 | two complexes of AmyB with carbohydrate ligands: the 1.35-A-resolution structure with the acarbose transglycosylation product, and the 2.2-A-resolution conplex of AmyB with alpha-cyclodextrin and hydrolysis products of maltoheptaose. The AmyB-acarbose complex includes residues 15-599, 1 acarbose molecule, 1 acarbose derived nonsaccharide, 2 glucose molecules, 4 calcium ions, 1 sodium ion, and 576 water molecules. The AmyB-maltoheptaose/cyclodextrin complex includes residues 14-599, 2 maltotetraose molecules, 1 alpha-cyclodextrin molecule, 7 calcium ions, 1 sodium ion, and 260 water molecules. The active site in AMyB is located at the C-terminal end of TIM barrel, with the residues Asp350, Glu380, and Asp447 as the catalytic residues | Halothermothrix orenii |
Protein Variants
| EC Number | Protein Variants | Comment | Organism |
|---|---|---|---|
| 3.2.1.1 | DELTAAmyB | lacking the N-domain, with no significant difference between the rates of soluble starch degradation, indicating that the N-domain does not play a direct role in catalysis with this substrate. For insoluble starch AmyB shows increase binding compared with DELTAAmyB, suggesting that the N-domain enhances the ability of AmyB to bind this substrate. The temperature stability of AmyB and DELTAAmyB, lacking the N-domain are strongly influenced by NaCl concentration, shown by an increasing melting temperature with increased NaCl concentrations up to 4-4.5 M | Halothermothrix orenii |
Localization
| EC Number | Localization | Comment | Organism | GeneOntology No. | Textmining |
|---|---|---|---|---|---|
| 3.2.1.1 | membrane | bound | Halothermothrix orenii | 16020 | - |
Metals/Ions
| EC Number | Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|---|
| 3.2.1.1 | Ca2+ | strictly dependent, retaining below 20% activity in the absence of CaCl2. The optimal concentration is ca. 0.2 mM | Halothermothrix orenii |  |
| 3.2.1.1 | NaCl | the enzyme is active over a broad range of salt concentrations, with optimum activity at 0.9 M. At 1.7, 2.6, and 4.3 M NaCl AmyB ist 80, 60, and 12% active, respectively. AmyB is a halophilic enzyme, but is still above 45% active in the absence of salt | Halothermothrix orenii | |
Molecular Weight [Da]
| EC Number | Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|---|
| 3.2.1.1 | 71000 | - |
excluding the lipoprotein signal peptide | Halothermothrix orenii |
Organism
| EC Number | Organism | UniProt | Comment | Textmining |
|---|---|---|---|---|
| 3.2.1.1 | Halothermothrix orenii | - |
- |
- |
Substrates and Products (Substrate)
| EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 3.2.1.1 | alpha-cyclodextrin + H2O | 1.1% relative activity compared to amylose as substrate, pH 8.0. 65°C | Halothermothrix orenii | ? | - |
? | |
| 3.2.1.1 | amylopectin + H2O | 74.6% relative activity compared to amylose as substrate, pH 8.0. 65°C | Halothermothrix orenii | ? | - |
? | |
| 3.2.1.1 | amylose + H2O | 100% relative activity as reference for substrate specificity studies, pH 8.0. 65°C | Halothermothrix orenii | ? | - |
? | |
| 3.2.1.1 | beta-cyclodextrin + H2O | 3.7% relative activity compared to amylose as substrate, pH 8.0. 65°C | Halothermothrix orenii | ? | - |
? | |
| 3.2.1.1 | gamma-cyclodextrin + H2O | 2.6% relative activity compared to amylose as substrate, pH 8.0. 65°C | Halothermothrix orenii | ? | - |
? | |
| 3.2.1.1 | glycogen + H2O | 16.7% relative activity compared to amylose as substrate, pH 8.0. 65°C | Halothermothrix orenii | ? | - |
? | |
| 3.2.1.1 | pullulan + H2O | 2.5% relative activity compared to amylose as substrate, pH 8.0. 65°C | Halothermothrix orenii | ? | - |
? | |
| 3.2.1.1 | starch + H2O | soluble starch, 93.9% relative activity compared to amylose as substrate, pH 8.0, 65°C | Halothermothrix orenii | ? | - |
? | |
Synonyms
| EC Number | Synonyms | Comment | Organism |
|---|---|---|---|
| 3.2.1.1 | AmyB | - |
Halothermothrix orenii |
Temperature Optimum [°C]
| EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 3.2.1.1 | 65 | - |
optimum and assay at | Halothermothrix orenii |
Temperature Range [°C]
| EC Number | Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 3.2.1.1 | 37 | 80 | retains above 40% activity within the range of 37-80°C, with an optimum at 65°C | Halothermothrix orenii |
Temperature Stability [°C]
| EC Number | Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 3.2.1.1 | additional information | - |
the temperature stability of AmyB and DELTAAmyB, lacking the N-domain are strongly influenced by NaCl concentration, shown by an increasing melting temperature with increased NaCl concentrations up to 4-4.5 M | Halothermothrix orenii |
| 3.2.1.1 | 60 | - |
fully active after preincubation at 60°C in the presence of soluble starch. In the absence of starch AmyB is inactivated rapidly after 15 min of preincubation | Halothermothrix orenii |
| 3.2.1.1 | 63 | - |
Tm value, the maximum thermal stability for AmyB and DELTAAmyB, lacking the N-domain are obtained at pH above 6.0 and above 3.0 M NaCl | Halothermothrix orenii |
| 3.2.1.1 | 80 | - |
after preincubation in the presence of starch, activity falls off linearly with time to reach 10% after 2 h, with a half-life of above 1 h at 80°C. In the absence of starch AmyB is inactivated rapidly after 15 min of preincubation | Halothermothrix orenii |
pH Optimum
| EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|---|
| 3.2.1.1 | 7 | - |
- |
Halothermothrix orenii |
| 3.2.1.1 | 8 | - |
assay at | Halothermothrix orenii |
pH Range
| EC Number | pH Minimum | pH Maximum | Comment | Organism |
|---|---|---|---|---|
| 3.2.1.1 | 5.2 | 9.3 | above 20% activity within the range of pH 5.2-9.3, with an optimum at pH 7.0 | Halothermothrix orenii |
pH Stability
| EC Number | pH Stability | pH Stability Maximum | Comment | Organism |
|---|---|---|---|---|
| 3.2.1.1 | 5 | - |
AmyB and the variant DELTAAmyB, lacking the N-domain depend only weakly on pH. At pH 5.0 both proteins are destabilized | Halothermothrix orenii |
| 3.2.1.1 | 7 | - |
AmyB and the variant DELTAAmyB, lacking the N-domain depend only weakly on pH. At pH 7.0 both proteins are equally stabilized | Halothermothrix orenii |
| 3.2.1.1 | 7.5 | 8.5 | AmyB and the variant DELTAAmyB, lacking the N-domain depend only weakly on pH. At 7.5-8.5 they show almost identical Tm values | Halothermothrix orenii |
pI Value
| EC Number | Organism | Comment | pI Value Maximum | pI Value |
|---|---|---|---|---|
| 3.2.1.1 | Halothermothrix orenii | theoretical value | - |
4.4 |
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