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Literature summary for 3.2.1.70 extracted from
- Enzymology and structure of the GH13_31 glucan 1,6-alpha-glucosidase that confers isomaltooligosaccharide utilization in the probiotic Lactobacillus acidophilus NCFM (2012), J. Bacteriol., 194, 4249-4259.
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| gene LBA0264, DNA and amino acid sequence determination and analysis, genetic organization, sequence comparisons and phylogenetic tree, the enzyme is encoded in the maltooligosaccharide operon, semiquantitative PCR expression analysis, cloning in Escherichia coli strain TOP10, expression of C-terminally His6-tagged enzyme in Escherichia coli strain BL21(DE3) | Lactobacillus acidophilus |
Crystallization (Commentary)
| Crystallization (Comment) | Organism |
|---|---|
| purified recombinant His-tagged enzyme, sitting drop vapour diffusion method, mixing of 16 mg/ml protein in 20 mM MES-NaOH, pH 6.5, 100 mM NaCl, and 2 mM CaCl2, with reservoir solution containing 20% glycerol, 16% PEG 8000, and 0.1 M MES, pH 6.5, in a 2:1 ratio, room temperature, X-ray diffraction structure determination and analysis at 2.05 A resolution, molecular replacement and modelling | Lactobacillus acidophilus |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| D-glucose | product inhibition | Lactobacillus acidophilus |  |
KM Value [mM]
| KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| additional information | - |
additional information | Michaelis-Menten kinetics | Lactobacillus acidophilus | |
| 2.38 | - |
4-nitrophenyl-alpha-D-glucopyranoside | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 3.9 | - |
panose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 14.1 | - |
isomaltotriose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 16.1 | - |
isomaltotetraose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 22.5 | - |
isomaltose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 29.1 | - |
Dextran | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Ca2+ | stabilizes, a Ca2 is bound in domain A in a loop located just before the first helix in a binding site, the Ca2+ octahedral coordination shell comprises the side chains of Asp20, Asn22, Asp24, and Asp28, the carbonyl oxygen of Ile26, and a water molecule | Lactobacillus acidophilus | |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| dextran + H2O | Lactobacillus acidophilus | - |
D-glucose + ? | - |
? | |
| dextran + H2O | Lactobacillus acidophilus NCFM | - |
D-glucose + ? | - |
? | |
| isomaltooligosaccharide + H2O | Lactobacillus acidophilus | degradation | D-glucose + ? | - |
? | |
| isomaltose + H2O | Lactobacillus acidophilus | - |
2 D-glucose | - |
? | |
| isomaltose + H2O | Lactobacillus acidophilus NCFM | - |
2 D-glucose | - |
? | |
| isomaltotetraose + H2O | Lactobacillus acidophilus | - |
? | - |
? | |
| isomaltotetraose + H2O | Lactobacillus acidophilus NCFM | - |
? | - |
? | |
| isomaltotriose + H2O | Lactobacillus acidophilus | - |
? | - |
? | |
| isomaltotriose + H2O | Lactobacillus acidophilus NCFM | - |
? | - |
? | |
| panose + H2O | Lactobacillus acidophilus | - |
D-glucose + ? | - |
? | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Lactobacillus acidophilus | Q5FMB7 | gene LBA0264 | - |
| Lactobacillus acidophilus NCFM | Q5FMB7 | gene LBA0264 | - |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant His6-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, anion exchange chromatography, and ultrafiltration, to homogeneity | Lactobacillus acidophilus |
Specific Activity [micromol/min/mg]
| Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|
| 335 | - |
purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| 4-nitrophenyl-alpha-D-glucopyranoside + H2O | - |
Lactobacillus acidophilus | 4-nitrophenol + D-glucose | - |
? | |
| dextran + H2O | - |
Lactobacillus acidophilus | D-glucose + ? | - |
? | |
| dextran + H2O | catalytic efficiency on dextran and dextran/panose | Lactobacillus acidophilus | D-glucose + ? | - |
? | |
| dextran + H2O | - |
Lactobacillus acidophilus NCFM | D-glucose + ? | - |
? | |
| dextran + H2O | catalytic efficiency on dextran and dextran/panose | Lactobacillus acidophilus NCFM | D-glucose + ? | - |
? | |
| isomaltooligosaccharide + H2O | - |
Lactobacillus acidophilus | D-glucose + ? | - |
? | |
| isomaltooligosaccharide + H2O | degradation | Lactobacillus acidophilus | D-glucose + ? | - |
? | |
| isomaltose + H2O | - |
Lactobacillus acidophilus | 2 D-glucose | - |
? | |
| isomaltose + H2O | - |
Lactobacillus acidophilus NCFM | 2 D-glucose | - |
? | |
| isomaltotetraose + H2O | - |
Lactobacillus acidophilus | ? | - |
? | |
| isomaltotetraose + H2O | - |
Lactobacillus acidophilus NCFM | ? | - |
? | |
| isomaltotriose + H2O | - |
Lactobacillus acidophilus | ? | - |
? | |
| isomaltotriose + H2O | - |
Lactobacillus acidophilus NCFM | ? | - |
? | |
| panose + H2O | - |
Lactobacillus acidophilus | D-glucose + ? | - |
? | |
| panose + H2O | catalytic efficiency on dextran/panose | Lactobacillus acidophilus | D-glucose + ? | - |
? | |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| GH13_31 glucan 1,6-alpha-glucosidase | - |
Lactobacillus acidophilus |
| LaGH13_31 | - |
Lactobacillus acidophilus |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 39 | - |
- |
Lactobacillus acidophilus |
Temperature Stability [°C]
| Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 37 | - |
purified recombinant His6-tagged enzyme, pH 6.0, inactivation rate constant is 0.0001/min, T1/2 at 37°C is 4.8 days | Lactobacillus acidophilus |
| 50 | - |
purified recombinant His6-tagged enzyme, pH 6.0, inactivation rate constant is 0.0152/min, T1/2 at 50°C is 46 min | Lactobacillus acidophilus |
Turnover Number [1/s]
| Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 384 | - |
Dextran | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 475 | - |
isomaltotriose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 501 | - |
isomaltotetraose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 517 | - |
isomaltose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 597 | - |
4-nitrophenyl-alpha-D-glucopyranoside | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 612 | - |
panose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 5.5 | - |
- |
Lactobacillus acidophilus |
pH Stability
| pH Stability | pH Stability Maximum | Comment | Organism |
|---|---|---|---|
| 4 | 8 | purified recombinant His6-tagged enzyme, over 50% activity remaining | Lactobacillus acidophilus |
Ki Value [mM]
| Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 40 | - |
D-glucose | pH 6.0, 37°C, recombinant enzyme | Lactobacillus acidophilus | |
pI Value
| Organism | Comment | pI Value Maximum | pI Value |
|---|---|---|---|
| Lactobacillus acidophilus | recombinant His-tagged enzyme, isoelectric focusing | - |
6.5 |
Expression
| Organism | Comment | Expression |
|---|---|---|
| Lactobacillus acidophilus | besides the isolated locus encoding a GH13_31 enzyme, the ABC transporter and another GH13 in the maltooligosaccharide operon are induced in response to IMO or maltotetraose suggesting coregulation of alpha-1,6- and alpha-1,4-glucooligosaccharide utilization loci in Lactobacillus acidophilus NCFM | additional information |
| Lactobacillus acidophilus | the enzyme is induced by isomaltooligosaccharides and maltotetraose | up |
General Information
| General Information | Comment | Organism |
|---|---|---|
| evolution | the enzyme belongs to the glycosyl hydrolase family 13 subfamily 31 | Lactobacillus acidophilus |
| additional information | phylogenetic and activity motif analysis, overview. Catalytic residues are Asp198 and Glu240, active site structure, overview | Lactobacillus acidophilus |
| physiological function | coregulation of alpha-1,6- and alpha-1,4-glucooligosaccharide utilization loci in the organism | Lactobacillus acidophilus |
kcat/KM [mM/s]
| kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|
| 13.6 | - |
Dextran | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 23 | - |
isomaltose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 31 | - |
isomaltotetraose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 34 | - |
isomaltotriose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 157 | - |
panose | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
| 251 | - |
4-nitrophenyl-alpha-D-glucopyranoside | purified recombinant enzyme, pH 6.0, 37°C | Lactobacillus acidophilus | |
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