Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary extracted from

  • Nakai, H.; Petersen, B.O.; Westphal, Y.; Dilokpimol, A.; Abou Hachem, M.; Duus, J.?.; Schols, H.A.; Svensson, B.
    Rational engineering of Lactobacillus acidophilus NCFM maltose phosphorylase into either trehalose or kojibiose dual specificity phosphorylase (2010), Protein Eng. Des. Sel., 23, 781-787.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.4.1.8 expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 (DE3) Lactobacillus acidophilus
2.4.1.230 expression of His-tagged LaMP loop 3 mutants in Escherichia coli strain BL21(DE3) Lactobacillus acidophilus

Protein Variants

EC Number Protein Variants Comment Organism
2.4.1.8 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
2.4.1.230 additional information substitution of maltose phosphorylase, EC 2.4.1.8, 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 1+, by corresponding to segments from Ser426-Ala431 in trehalose phosphorylase, EC 2.4.1.64, and Thr419-Phe427 in kojibiose phosphorylase, EC 2.4.1.230, thus confers the maltose phosphorylase with phosphorolytic activity towards trehalose and kojibiose, respectively Lactobacillus acidophilus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.4.1.230 9.3
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 3 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.230 16
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 2 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.230 84
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 1 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.231 22
-
 alpha,alpha-trehalose loop 3 variant TP3, in 40 mM phosphate-citrate pH 6.2 at 45°C Lactobacillus acidophilus
2.4.1.231 53
-
 alpha,alpha-trehalose loop 3 variant TP2, in 40 mM phosphate-citrate pH 6.2 at 45°C Lactobacillus acidophilus

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.4.1.8 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
2.4.1.8 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
2.4.1.230 2-alpha-D-glucosyl-D-glucose + phosphate Thermoanaerobacter brockii
-
 D-glucose + beta-D-glucose 1-phosphate
-
 ?
2.4.1.230 2-alpha-D-glucosyl-D-glucose + phosphate Thermoanaerobacter brockii ATCC 35047
-
 D-glucose + beta-D-glucose 1-phosphate
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.4.1.8 Lactobacillus acidophilus Q5FI04
-
-
2.4.1.8 Lactobacillus acidophilus NCFM Q5FI04
-
-
2.4.1.230 Lactobacillus acidophilus
-
-
-
2.4.1.230 Thermoanaerobacter brockii
-
-
-
2.4.1.230 Thermoanaerobacter brockii ATCC 35047
-
-
-
2.4.1.231 Lactobacillus acidophilus
-
-
-
2.4.1.231 Lactobacillus acidophilus NCFM
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.4.1.8 recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 (DE3) by affinity chromatography and dialysis Lactobacillus acidophilus
2.4.1.230 recombinant His-tagged LaMP loop 3 mutants from Escherichia coli strain BL21(DE3) by affinity chromatography and dialysis Lactobacillus acidophilus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.4.1.8 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
2.4.1.8 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
2.4.1.8 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
2.4.1.8 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
2.4.1.8 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 ?
-
 ?
2.4.1.8 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 ?
-
 ?
2.4.1.230 2-alpha-D-glucosyl-D-glucose + phosphate
-
 Thermoanaerobacter brockii D-glucose + beta-D-glucose 1-phosphate
-
 ?
2.4.1.230 2-alpha-D-glucosyl-D-glucose + phosphate activity of a LaMP loop 3 maltose phosphorylase mutant, no activity with wild-type maltose phosphorylase, which is strictly specific for maltose Lactobacillus acidophilus D-glucose + beta-D-glucose 1-phosphate
-
 r
2.4.1.230 2-alpha-D-glucosyl-D-glucose + phosphate activity of a LaMP loop 3 maltose phosphorylase mutant, no activity with wild-type maltose phosphorylase, which is strictly specific for maltose Lactobacillus acidophilus NCFM D-glucose + beta-D-glucose 1-phosphate
-
 r
2.4.1.230 2-alpha-D-glucosyl-D-glucose + phosphate
-
 Thermoanaerobacter brockii ATCC 35047 D-glucose + beta-D-glucose 1-phosphate
-
 ?
2.4.1.231 alpha,alpha-trehalose + phosphate
-
 Lactobacillus acidophilus alpha-D-glucose + alpha-D-glucose 1-phosphate
-
 ?
2.4.1.231 alpha,alpha-trehalose + phosphate
-
 Lactobacillus acidophilus NCFM alpha-D-glucose + alpha-D-glucose 1-phosphate
-
 ?
2.4.1.231 additional information the maltose phosphorylase loop 3 variants TP2 and TP3 are capable of phosphorolysing trehalose (alpha, alpha1) as well as maltose (alpha-1,4),but show no activity towards kojibiose (alpha-1,2), nigerose (alpha-1,3) or isomaltose (alpha-1,6). The catalytic efficiency of TP3 towards trehalose is 19fold higher than of TP2 Lactobacillus acidophilus ?
-
 ?
2.4.1.231 additional information the maltose phosphorylase loop 3 variants TP2 and TP3 are capable of phosphorolysing trehalose (alpha, alpha1) as well as maltose (alpha-1,4),but show no activity towards kojibiose (alpha-1,2), nigerose (alpha-1,3) or isomaltose (alpha-1,6). The catalytic efficiency of TP3 towards trehalose is 19fold higher than of TP2 Lactobacillus acidophilus NCFM ?
-
 ?

Subunits

EC Number Subunits Comment Organism
2.4.1.8 More sequence compariosn with GH65 trehalose phosphorylase and kojibiose phosphorylase, and homology modelling, overview Lactobacillus acidophilus

Synonyms

EC Number Synonyms Comment Organism
2.4.1.230 kojibiose phosphorylase
-
 Thermoanaerobacter brockii
2.4.1.231 TP2 a loop 3 variant of Lactobacillus acidophilus maltose phosphorylase Lactobacillus acidophilus
2.4.1.231 TP3 a loop 3 variant of Lactobacillus acidophilus maltose phosphorylase Lactobacillus acidophilus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.4.1.8 45
-
 phosphorolysis Lactobacillus acidophilus
2.4.1.230 45
-
 phosphorolysis and reverse phosphorolysis Lactobacillus acidophilus
2.4.1.231 45 50
-
 Lactobacillus acidophilus

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
2.4.1.8 90
-
 purified recombinant enzyme, 15 min, inactivation Lactobacillus acidophilus
2.4.1.231 45 50
-
 Lactobacillus acidophilus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.4.1.230 0.097
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 1 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.230 0.12
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 2 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.230 2.4
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 3 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.231 0.33
-
 alpha,alpha-trehalose loop 3 variant TP2, in 40 mM phosphate-citrate pH 6.2 at 45°C Lactobacillus acidophilus
2.4.1.231 2.7
-
 alpha,alpha-trehalose loop 3 variant TP3, in 40 mM phosphate-citrate pH 6.2 at 45°C Lactobacillus acidophilus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.4.1.8 6.2
-
 phosphorolysis Lactobacillus acidophilus
2.4.1.230 3.4
-
 reverse phosphorolysis Lactobacillus acidophilus
2.4.1.230 6.2
-
 phosphorolysis Lactobacillus acidophilus
2.4.1.231 6.3 6.7
-
 Lactobacillus acidophilus

pH Stability

EC Number pH Stability pH Stability Maximum Comment Organism
2.4.1.231 3.2 8.9
-
 Lactobacillus acidophilus

General Information

EC Number General Information Comment Organism
2.4.1.8 evolution NCFM maltose phosphorylase belongs to the (alpha/alpha)6-barrel glycoside hydrolase family 65, GH65 Lactobacillus acidophilus
2.4.1.8 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
2.4.1.230 evolution the enzyme is a member of glycoside hydrolase family 65, GH65 Lactobacillus acidophilus
2.4.1.230 evolution the enzyme is a member of glycoside hydrolase family 65, GH65 Thermoanaerobacter brockii
2.4.1.230 additional information structure-function relationship analysis, homology modelling, overview. Substitution of maltose phosphorylase, EC 2.4.1.8, 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 1+, by corresponding to segments from Ser426-Ala431 in trehalose phosphorylase, EC 2.4.1.64, and Thr419-Phe427 in kojibiose phosphorylase, EC 2.4.1.230, thus confers the maltose phosphorylase with phosphorolytic activity towards trehalose and kojibiose, respectively. The loop 3 in GH65 disaccharide phosphorylase is a key determinant for specificity both in phosphorolysis and in regiospecific reverse phosphorolysis Lactobacillus acidophilus
2.4.1.230 additional information structure-function relationship analysis, overview. The sequence Thr419-Phe427 is responsible for substrate specificity of the enzyme for kojibiose. The loop 3 in GH65 disaccharide phosphorylase is a key determinant for specificity both in phosphorolysis and in regiospecific reverse phosphorolysis Thermoanaerobacter brockii

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.4.1.230 0.0011
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 1 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.230 0.0075
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 2 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.230 0.26
-
 2-alpha-D-glucosyl-D-glucose pH 6.2, 45°C, recombinant loop 3 mutant 3 of maltose phosphorylase Lactobacillus acidophilus
2.4.1.231 0.0062
-
 alpha,alpha-trehalose loop 3 variant TP2, in 40 mM phosphate-citrate pH 6.2 at 45°C Lactobacillus acidophilus
2.4.1.231 0.12
-
 alpha,alpha-trehalose loop 3 variant TP3, in 40 mM phosphate-citrate pH 6.2 at 45°C Lactobacillus acidophilus