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Literature summary for 2.4.1.8 extracted from
- Rational engineering of Lactobacillus acidophilus NCFM maltose phosphorylase into either trehalose or kojibiose dual specificity phosphorylase (2010), Protein Eng. Des. Sel., 23, 781-787.
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 (DE3) | Lactobacillus acidophilus |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| additional information | the (alpha/alpha)6-barrel loop 3 is a target for specificity engineering to achieve formation of trehalose and kojibiose by the enzyme in yields superior of maltose by reverse phosphorolysis with (alpha1, alpha1)- and alpha-(1,2)-regioselectivity, respectively. Substitution of LaMP His413-Glu421, His413-Ile418 and His413-Glu415 from loop 3, that include His413 and Glu415 presumably recognising the alpha-anomeric O-1 group of the glucose moiety at subsite 11, by corresponding segments from Ser426-Ala431 in trehalose phosphorylase and Thr419-Phe427 in kojibiose phosphorylase, thus confers LaMP with phosphorolytic activity towards trehalose and kojibiose, respectively. Substrate specificity of LaMP loop 3 variants in phosphorolysis, overview | Lactobacillus acidophilus |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| maltose + phosphate | Lactobacillus acidophilus | the enzyme catalyzes both phosphorolysis of maltose and formation of maltose by reverse phosphorolysis with beta-glucose 1-phosphate and glucose as donor and acceptor, respectively | D-glucose + beta-D-glucose 1-phosphate | - |
r |  |
| maltose + phosphate | Lactobacillus acidophilus NCFM | the enzyme catalyzes both phosphorolysis of maltose and formation of maltose by reverse phosphorolysis with beta-glucose 1-phosphate and glucose as donor and acceptor, respectively | D-glucose + beta-D-glucose 1-phosphate | - |
r | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Lactobacillus acidophilus | Q5FI04 | - |
- |
| Lactobacillus acidophilus NCFM | Q5FI04 | - |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 (DE3) by affinity chromatography and dialysis | Lactobacillus acidophilus |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| maltose + phosphate | the enzyme catalyzes both phosphorolysis of maltose and formation of maltose by reverse phosphorolysis with beta-glucose 1-phosphate and glucose as donor and acceptor, respectively | Lactobacillus acidophilus | D-glucose + beta-D-glucose 1-phosphate | - |
r | |
| maltose + phosphate | the enzyme catalyzes both phosphorolysis of maltose and formation of maltose by reverse phosphorolysis with beta-glucose 1-phosphate and glucose as donor and acceptor, respectively. Maltose phosphorylase catalyses phosphorolysis of maltose with inversion of the anomeric configuration to release beta-glucose 1-phosphate and glucose | Lactobacillus acidophilus | D-glucose + beta-D-glucose 1-phosphate | - |
r | |
| maltose + phosphate | the enzyme catalyzes both phosphorolysis of maltose and formation of maltose by reverse phosphorolysis with beta-glucose 1-phosphate and glucose as donor and acceptor, respectively | Lactobacillus acidophilus NCFM | D-glucose + beta-D-glucose 1-phosphate | - |
r | |
| maltose + phosphate | the enzyme catalyzes both phosphorolysis of maltose and formation of maltose by reverse phosphorolysis with beta-glucose 1-phosphate and glucose as donor and acceptor, respectively. Maltose phosphorylase catalyses phosphorolysis of maltose with inversion of the anomeric configuration to release beta-glucose 1-phosphate and glucose | Lactobacillus acidophilus NCFM | D-glucose + beta-D-glucose 1-phosphate | - |
r | |
| additional information | substrate specificity, overview. The wild-type enzyme does not show activity with trehalose, kojibiose, nigerose, and isomaltose, while its loop 3 mutants do to different degrees, overview | Lactobacillus acidophilus | ? | - |
? | |
| additional information | substrate specificity, overview. The wild-type enzyme does not show activity with trehalose, kojibiose, nigerose, and isomaltose, while its loop 3 mutants do to different degrees, overview | Lactobacillus acidophilus NCFM | ? | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| More | sequence compariosn with GH65 trehalose phosphorylase and kojibiose phosphorylase, and homology modelling, overview | Lactobacillus acidophilus |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 45 | - |
phosphorolysis | Lactobacillus acidophilus |
Temperature Stability [°C]
| Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 90 | - |
purified recombinant enzyme, 15 min, inactivation | Lactobacillus acidophilus |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 6.2 | - |
phosphorolysis | Lactobacillus acidophilus |
General Information
| General Information | Comment | Organism |
|---|---|---|
| evolution | NCFM maltose phosphorylase belongs to the (alpha/alpha)6-barrel glycoside hydrolase family 65, GH65 | Lactobacillus acidophilus |
| additional information | the (alpha/alpha)6-barrel loop 3 forms the rim of the active site pocket and is a key determinant for specificity both in phosphorolysis and in regiospecific reverse phosphorolysis | Lactobacillus acidophilus |
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