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Literature summary for 3.2.1.1 extracted from

  • Bozonnet, S.; Jensen, M.T.; Nielsen, M.M.; Aghajari, N.; Jensen, M.H.; Kramhoft, B.; Willemoes, M.; Tranier, S.; Haser, R.; Svensson, B.
    The pair of sugar tongs site on the non-catalytic domain C of barley alpha-amylase participates in substrate binding and activity (2007), FEBS J., 274, 5055-5067.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expression in Pichia pastoris Hordeum vulgare

Crystallization (Commentary)

Crystallization (Comment) Organism
the structures of AMY1 Y380A and S378P are compared with the wild-type enzyme both in free form and in complex with acarbose Hordeum vulgare

Protein Variants

Protein Variants Comment Organism
S378P kcat/KM for amylose is 1.2fold lower than wild-type value. kcat/KM for 2-chloro-4-nitrophenyl beta-D-maltoheptaoside is nearly identical to wild-type value Hordeum vulgare
S378T kcat/KM for amylose is 1.4fold lower than wild-type value. kcat/KM for 2-chloro-4-nitrophenyl beta-D-maltoheptaoside is nearly identical to wild-type value Hordeum vulgare
Y380A mutant fails to bind to beta-cyclodextrin-Sepharose, a starch-mimic resin used for alpha-amylase affinity purification. The Kd for beta-cyclodextrin binding to Y380A is 1.4 mm compared to 0.20-0.25 mM for the wild-type, S378P and S378T enzymes. Crystal structures of both wild-type and S378P enzymes, but not the Y380A enzyme, shows binding of the pseudotetrasaccharide acarbose at the sugar tongs site. beta-Cyclodextrin both inhibits binding to and suppresses activity on starch granules for wild-type and S378P enzymes, but does not affect these properties of Y380A. Y380A enzyme hydrolyzes amylose with reduced multiple attack. kcat/KM for amylose is 3.7fold lower than wild-type value. kcat/KM for 2-chloro-4-nitrophenyl beta-D-maltoheptaoside is 2.4fold lower than wild-type value Hordeum vulgare
Y380F kcat/KM for amylose is fold lowerthan wild-type value. kcat/KM for is 2.6fold lower than wild-type value. kcat/KM for 2-chloro-4-nitrophenyl beta-D-maltoheptaoside is 1.1fold higher than wild-type value Hordeum vulgare
Y380M mutant fails to bind to beta-cyclodextrin-Sepharose, a starch-mimic resin used for alpha-amylase affinity purification. The Kd for beta-cyclodextrin binding to Y380M is 1.4 mm compared to 0.20-0.25 mM for the wild-type, S378P and S378T enzymes. kcat/KM for amylose is 2.3fold lower than wild-type value. kcat/KM for 2-chloro-4-nitrophenyl beta-D-maltoheptaoside is 1.8fold lower than wild-type value Hordeum vulgare

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information KM-values for amylose: 0.363 mg/ml for mutant enzyme Y380A, 0.351 mg/ml for mutant enzyme Y380M, 0.391 mg/ml for mutant enzyme Y380F, 0.203 mg/ml for mutant enzyme S378P, 0.208 mg/ml for mutant enzyme S378T, 0.19 mg/ml for alpha-amylase 1 Hordeum vulgare
0.669
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme Y380A Hordeum vulgare
0.724
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme Y380F Hordeum vulgare
0.735
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme S378T Hordeum vulgare
0.758
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside alpha-amylase 1 Hordeum vulgare
0.861
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme S378P Hordeum vulgare
0.871
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme Y380M Hordeum vulgare

Organism

Organism UniProt Comment Textmining
Hordeum vulgare P00693
-
-

Purification (Commentary)

Purification (Comment) Organism
-
Hordeum vulgare

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2-chloro-4-nitrophenyl beta-D-maltoheptaoside + H2O
-
Hordeum vulgare ?
-
?
amylose DP440 + H2O
-
Hordeum vulgare ?
-
?

Synonyms

Synonyms Comment Organism
alpha-amylase type A isozyme
-
Hordeum vulgare
alpha-amylases 1
-
Hordeum vulgare
AMY1
-
Hordeum vulgare
Low pI alpha-amylase
-
Hordeum vulgare

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
19
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme Y380A Hordeum vulgare
34
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme Y380M Hordeum vulgare
48
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme S378T Hordeum vulgare
52
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside alpha-amylase 1 Hordeum vulgare
56
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme Y380F Hordeum vulgare
59
-
2-chloro-4-nitrophenyl beta-D-maltoheptaoside mutant enzyme S378P Hordeum vulgare
95
-
amylose DP440 mutant enzyme Y380A Hordeum vulgare
144
-
amylose DP440 mutant enzyme S378T Hordeum vulgare
149
-
amylose DP440 mutant enzyme Y380M Hordeum vulgare
162
-
amylose DP440 mutant enzyme Y380F Hordeum vulgare
163
-
amylose DP440 mutant enzyme S378P Hordeum vulgare
185
-
amylose DP440 alpha-amylase 1 Hordeum vulgare