Cloned (Comment) | Organism |
---|---|
expression in Pichia pastoris | Hordeum vulgare |
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 | 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 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 | UniProt | Comment | Textmining |
---|---|---|---|
Hordeum vulgare | P00693 | - |
- |
Purification (Comment) | Organism |
---|---|
- |
Hordeum vulgare |
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 | 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 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 |