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Literature summary for 3.2.1.2 extracted from

  • Ye, Z.; Miyake, H.; Tatsumi, M.; Nishimura, S.; Nitta, Y.
    Two additional carbohydrate-binding sites of beta-amylase from Bacillus cereus var. mycoides are involved in hydrolysis and raw starch-binding (2004), J. Biochem., 135, 355-363.
    View publication on PubMed
Search for more literature for 3.2.1.2

Cloned(Commentary)

Cloned (Comment) Organism
wild-type and mutant enzymes, expression in Escherichia coli BL21(DE3) Bacillus cereus

Crystallization (Commentary)

Crystallization (Comment) Organism
-
 Bacillus cereus

Protein Variants

Protein Variants Comment Organism
additional information mutation of two of the three carbohydrate-binding sites aside from the active site: Site2 in domain B and Site1 in domain C Bacillus cereus
S235A binding parameters to raw corn starch, kinetic parameters for the hydrolysis of amylose, 88% of wild-type activity with soluble starch as substrate Bacillus cereus
S235A/Y249A double mutant, binding parameters to raw corn starch, kinetic parameters for the hydrolysis of amylose, 63% of wild-type activity with soluble starch as substrate Bacillus cereus
S235A/Y249A/W449F/W495F quadruple mutant, kinetic parameters for the hydrolysis of amylose, 51% of wild-type activity with soluble starch as substrate Bacillus cereus
W449F binding parameters to raw corn starch Bacillus cereus
W449F/W495F double mutant, binding parameters to raw corn starch, kinetic parameters for the hydrolysis of amylose, 61% of wild-type activity with soluble starch as substrate Bacillus cereus
W495F binding parameters to raw corn starch Bacillus cereus
W51F beta-amylase mutant Bacillus cereus
Y249A binding parameters to raw corn starch, kinetic parameters for the hydrolysis of amylose, 80% of wild-type activity with soluble starch as substrate Bacillus cereus

Inhibitors

Inhibitors Comment Organism Structure
cyclohexaamylose alpha-CD, competitive inhibitor Bacillus cereus
maltitol behaves as a mixed-type or competitive inhibitor depending on the chain length of the substrate, inhibition mechanism, binds to Site2 in domain B and forms an abortive ESI complex when amylose is used as substrate Bacillus cereus
O-alpha-D-xylopyranosyl(1-4)O-alpha-D-glucopyranosyl(1-4)O-alpha-D-glucopyranoside mixed-type inhibition, two molecules bind to enzyme Bacillus cereus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information binding parameters of wild-type and mutant enzymes to raw corn starch Bacillus cereus
0.6
-
 amylose pH 7, 25°C, DPn: 16, S235A/Y249A/W449F/W495F quadruple mutant Bacillus cereus
0.61
-
 amylose pH 7, 25°C, DPn: 16, W449F/W495F double mutant Bacillus cereus
0.65
-
 amylose pH 7, 25°C, DPn: 16, S235A/Y249A double mutant Bacillus cereus
0.7
-
 amylose pH 7, 25°C, DPn: 16, S235A mutant Bacillus cereus
0.71
-
 amylose pH 7, 25°C, DPn: 16, Y249A mutant Bacillus cereus
0.72
-
 amylose pH 7, 25°C, DPn: 16, wild-type enzyme Bacillus cereus

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
58300
-
 x * 58300, Western blot analysis Bacillus cereus

Organism

Organism UniProt Comment Textmining
Bacillus cereus P36924 var. mycoides
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes Bacillus cereus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
amylose + H2O DPn is 16 Bacillus cereus ?
-
 ?
maltopentaose + H2O binding mode of substrate in the active site Bacillus cereus ?
-
 ?
additional information no hydrolysis of alpha-1,6-glucosidic linkages Bacillus cereus ?
-
 ?
starch + H2O catalyzes the hydrolysis of alpha-1,4-glucosidic linkages of soluble starch, and liberates beta-anomeric maltose from the nonreducing ends, exo-acting enzyme, composed of two functional domains, a catalytic domain: domains A and B, and starch-binding domain: domain C, beta-amylase has three carbohydrate-binding sites aside from the active site: two in domain B named Site2 and Site3, one in domain C named Site1, roles of these sites in the catalytic reaction and raw starch-binding, beta-amylase hardly hydrolyzes raw starch from wheat, corn, potato or sweet potato, but binds to it strongly Bacillus cereus ?
-
 ?

Subunits

Subunits Comment Organism
? x * 58300, Western blot analysis Bacillus cereus
More enzyme is composed of two functional domains, a catalytic domain: domains A and B, and starch-binding domain: domain C, beta-amylase has three carbohydrate-binding sites aside from the active site: two in domain B named Site2 and Site3, one in domain C named Site1, roles of these sites in the catalytic reaction and raw starch-binding Bacillus cereus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
 assay at Bacillus cereus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
additional information
-
 additional information
-
 Bacillus cereus

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
 assay at Bacillus cereus

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.45
-
 O-alpha-D-xylopyranosyl(1-4)O-alpha-D-glucopyranosyl(1-4)O-alpha-D-glucopyranoside pH 7, 25°C, wild-type enzyme, amylose as substrate Bacillus cereus
0.8
-
 cyclohexaamylose pH 7, 25°C, wild-type enzyme, amylose as substrate Bacillus cereus
2.8
-
 maltitol pH 7, 25°C, wild-type enzyme, amylose as substrate Bacillus cereus
3.1
-
 maltitol pH 7, 25°C, wild-type enzyme, maltopentaose as substrate Bacillus cereus
5.1
-
 maltitol pH 7, 25°C, S235A/Y249A double mutant, amylose as substrate Bacillus cereus
5.7
-
 maltitol pH 7, 25°C, S235A/Y249A/W449F/W495F quadruple mutant, amylose as substrate Bacillus cereus